Template:SDU-Denmark/js/app
// default/src/index.jsx $fsx.f[217] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const ReactDOM = $fsx.r(7); const Application_1 = $fsx.r(212); const $ = $fsx.r(102); $fsx.r(213); $fsx.r(214); $fsx.r(215); $fsx.r(216); let favicon = $('[rel="shortcut icon"]'); if (favicon.length) {
favicon[0].href = "";
} ReactDOM.render(React.createElement(Application_1.Application, null), document.getElementById("root")); //# sourceMappingURL=index.js.map } // default/src/components/application/Application.jsx $fsx.f[212] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const react_router_dom_1 = $fsx.r(29); const react_transition_group_1 = $fsx.r(43); const _ = $fsx.r(44); const ScrollToTop_1 = $fsx.r(45); const SubwayMenu_1 = $fsx.r(78); const react_scroll_progress_bar_1 = $fsx.r(79); const sitemap_1 = $fsx.r(209); $fsx.r(210); const timelines_1 = $fsx.r(211); const isMobile_1 = $fsx.r(58); const react_pose_1 = $fsx.r(57); const renderMergedProps = (component, ...rest) => {
const finalProps = Object.assign({}, ...rest);
return (React.createElement(component, finalProps));
}; const PropsRoute = (_a) => {
var { component } = _a, rest = tslib_1.__rest(_a, ["component"]);
return (React.createElement(react_router_dom_1.Route, Object.assign({}, rest, { render: routeProps => {
return renderMergedProps(component, routeProps, rest);
} })));
}; const VisibleOnScroll = react_pose_1.default.div({
hidden: {
top: "-210px",
transition: {
duration: 300,
ease: "easeOut"
}
},
visible: {
top: "0",
transition: {
duration: 300,
ease: "easeOut",
delay: 100
}
}
}); const OpenMenu = react_pose_1.default.div({
hidden: {
top: "200px",
transition: {
duration: 400,
ease: "easeOut",
delay: 0
}
},
visible: { top: "-100px" }
}); const MainContent = react_pose_1.default.div({
large: {
marginTop: "0"
},
small: {
marginTop: (el) => {
return "128px";
}
}
}); const updateCarousel = () => {
let el = document.querySelector(".slider-slide");
if (!el)
return;
let width = el.getBoundingClientRect().width;
if (width < 100) {
document.dispatchEvent(new Event("readystatechange"));
setTimeout(updateCarousel, 200);
}
else {
}
}; class Application extends React.Component {
constructor(props) {
super(props);
this.handleScroll = () => {
if (!this.subwayRef.current)
return;
if (document.body.classList.contains("page-tinder"))
return;
this.subwayRef.current.classList.remove('hidden-sub');
let menuHeight = this.subwayRef.current.getBoundingClientRect().height;
const { prevScrollpos } = this.state;
let currentScrollPos = window.pageYOffset;
const visible = (prevScrollpos > currentScrollPos) || (window.pageYOffset < menuHeight + 300);
if ((currentScrollPos - prevScrollpos) > 100 || (prevScrollpos - currentScrollPos) > 350) {
this.setState({
prevScrollpos: currentScrollPos,
isVisible: visible
});
}
else {
currentScrollPos = window.pageYOffset;
}
};
this.state = {
isVisible: true,
prevScrollpos: window.pageYOffset,
subwayIsLarge: true
};
this.subwayRef = React.createRef();
}
componentDidMount() {
window.addEventListener("scroll", _.throttle(this.handleScroll, 300));
}
componentWillUnmount() {
window.removeEventListener("scroll", this.handleScroll);
}
updateContentPosition(isLarge) {
if (this.state.subwayIsLarge !== isLarge) {
this.setState({
subwayIsLarge: isLarge
});
}
}
render() {
return (React.createElement("div", { className: `${isMobile_1.default ? "Application is-mobile" : "Application"}` },
isMobile_1.default &&
React.createElement("div", { id: "mobile-progress-bar" },
React.createElement(react_scroll_progress_bar_1.default, { height: "6", bgcolor: "#1DB2B0", duration: "0.2" })),
React.createElement(react_router_dom_1.BrowserRouter, { basename: "/Team:SDU-Denmark" },
React.createElement("div", { className: "top" },
React.createElement("div", { className: "logo" })),
React.createElement(VisibleOnScroll, { id: "subway", ref: this.subwayRef, pose: this.state.isVisible ? 'visible' : 'hidden' },
React.createElement(SubwayMenu_1.SubwayMenu, Object.assign({}, { sitemap: sitemap_1.default, flatSitemap: sitemap_1.FlatSitemap }, { onResizeMenu: this.updateContentPosition.bind(this) })),
React.createElement(OpenMenu, { onClick: () => {
this.setState({
isVisible: true
});
}, pose: this.state.isVisible ? 'hidden' : 'visible', className: "open-menu noselect" }, "Open")),
React.createElement(MainContent, { id: "main-content", pose: this.state.subwayIsLarge ? 'large' : 'small', style: {
overflowX: isMobile_1.default ? "hidden" : "visible"
} },
React.createElement(react_router_dom_1.Route, { render: ({ location }) => {
const { pathname, key } = location;
return (React.createElement(react_transition_group_1.TransitionGroup, { component: null },
React.createElement(react_transition_group_1.Transition, { key: key, appear: true, onEnter: (node, appears) => {
if (pathname == "/" || pathname == "") {
_.defer(() => {
this.setState({
isVisible: false
});
});
}
else {
_.defer(() => {
this.setState({
isVisible: true
});
});
}
timelines_1.play(pathname, node, appears);
setTimeout(() => {
updateCarousel();
}, 200);
}, onExit: ((node, appears) => timelines_1.exit(node)), timeout: { enter: 750, exit: 150 } },
React.createElement(React.Fragment, null,
React.createElement(ScrollToTop_1.default, null,
React.createElement(react_router_dom_1.Switch, { location: location }, sitemap_1.FlatSitemap.map((item, index) => {
let nextStation = sitemap_1.FlatSitemap.length > index ? sitemap_1.FlatSitemap[index + 1] : sitemap_1.FlatSitemap[0];
return React.createElement(PropsRoute, { exact: true, key: index, path: item.path, component: item.comp, nextStation: nextStation, title: item.displayName, nextChapter: false });
})))))));
} })))));
}
} exports.Application = Application; //# sourceMappingURL=Application.js.map } // default/src/components/application/ScrollToTop.jsx $fsx.f[45] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const react_router_1 = $fsx.r(28); class ScrollToTop extends React.Component {
componentDidUpdate(prevProps) {
if (this.props.location !== prevProps.location) {
window.scrollTo(0, 0);
}
}
render() {
return this.props.children;
}
} exports.default = react_router_1.withRouter(ScrollToTop); //# sourceMappingURL=ScrollToTop.js.map } // default/src/components/application/SubwayMenu.jsx $fsx.f[78] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const react_router_1 = $fsx.r(28); const _ = $fsx.r(44); const react_pose_1 = $fsx.r(57); const isMobile_1 = $fsx.r(58); const react_burger_menu_1 = $fsx.r(76); const STRIPE_WIDTH = 80; let styles = {
bmBurgerButton: {
position: 'fixed',
width: '26px',
height: '20px',
left: '26px',
top: '36px'
},
bmBurgerBars: {
background: '#193443'
},
bmBurgerBarsHover: {
background: '#3E6D8F'
},
bmCrossButton: {
height: '24px',
width: '24px'
},
bmCross: {
background: 'white',
},
bmMenuWrap: {
position: 'fixed',
height: '100%'
},
bmMenu: {
background: '#345B78',
padding: '2.5em 1.5em 0',
fontSize: '1.15em'
},
bmMorphShape: {
fill: '#345B78'
},
bmItemList: {
color: '#b8b7ad',
padding: '0.8em'
},
bmItem: {
display: 'inline-block'
},
bmOverlay: {
background: 'rgba(0, 0, 0, 0.3)'
}
}; const Menu = react_pose_1.default.div({
enlarge: {
paddingBottom: "40px",
transition: {
duration: 100,
ease: 'easeOut'
}
},
minimize: {
paddingBottom: "0px",
transition: {
duration: 100,
delay: 200,
ease: 'easeOut'
}
}
}); const MenuItem = react_pose_1.default.div({
hoverable: true,
pressable: true,
press: { scale: 0.9 },
init: {
scale: 1
},
hover: {
scale: 1.1
}
}); const SubMenuContainer = react_pose_1.default.div({
closedSubMenu: {
width: "0%",
opacity: 0,
transition: {
duration: 100,
width: { duration: 500 },
opacity: { duration: 300 },
}
},
openSubMenu: {
width: (arg) => {
let childrenWidth = _.chain(arg.element.children).map((child) => {
if (_.some(child.classList, (t) => t == "sub-item")) {
return child.getBoundingClientRect().width;
}
if (_.some(child.classList, (t) => t == "stripe")) {
return STRIPE_WIDTH;
}
return 0;
}).reduce((sum, child) => sum + child).value();
childrenWidth += 44;
let widthPercentage = (childrenWidth / arg.element.parentElement.getBoundingClientRect().width) * 100;
return `${Math.round(widthPercentage)}%`;
},
opacity: 1,
transition: {
width: {
duration: 400
},
opacity: { type: "spring", duration: 500 },
delay: 200
}
}
}); const SubMenuItem = react_pose_1.default.div({
hoverable: true,
pressable: true,
press: { scale: 0.9 },
init: {
scale: 1
},
hover: {
scale: 1.1
},
normal: { width: "auto" },
expanded: { width: "auto" }
}); const SubMenuItemBall = react_pose_1.default.div({
closedSubMenu: {
width: "1.0rem",
height: "1.0rem",
transition: {
duration: 300,
ease: "easeOut"
}
},
openSubMenu: {
width: "1.0rem",
height: "1.0rem",
transition: {
duration: 300,
ease: "easeOut"
}
}
}); class MenuComponent extends React.Component {
componentDidMount() {
setTimeout(() => {
this.setState({ isVisible: !this.state.isVisible });
}, 200);
}
constructor(props) {
super(props);
this.state = {
openSubMenu: false,
isVisible: false,
mobileMenuOpen: false
};
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(77);
});
}
handleStateChange(state) {
window.document.body.style.overflowY = state.isOpen ? "hidden" : "visible";
this.setState({ mobileMenuOpen: state.isOpen });
}
redirect(item) {
if (isMobile_1.default) {
this.setState({
mobileMenuOpen: false,
});
}
this.props.history.push(item);
}
isPredecessor(item) {
let props = this.props;
let currentIndex = _.findIndex(props.flatSitemap, (page, index) => {
return page.path == props.location.pathname;
});
let itemIndex = _.findIndex(props.flatSitemap, (page, index) => {
return page.path == item.path;
});
return itemIndex < currentIndex;
}
render() {
let props = this.props;
let activeClass = "active";
const { isVisible } = this.state;
let it = _.find(props.sitemap, (item) => props.location.pathname == item.path);
let isLarge = (it && it.children) || (_.find(props.sitemap, (ite) => {
if (!ite.children)
return;
return _.find(ite.children, (subIte) => props.location.pathname == subIte.path);
}));
this.props.onResizeMenu(!!isLarge);
const { openSubMenu } = this.state;
return (React.createElement(React.Fragment, null,
isMobile_1.default &&
React.createElement(react_burger_menu_1.bubble, { styles: styles, width: '100%', id: "mobile-menu", isOpen: this.state.mobileMenuOpen, onStateChange: (state) => this.handleStateChange(state) }, props.sitemap.map((item, index) => {
return (React.createElement(React.Fragment, { key: index },
React.createElement("div", { onClick: () => {
this.redirect(item.path);
}, className: (props.location.pathname == item.path) ? activeClass + " item" : "item" }, item.displayName),
item.children && item.children.length &&
React.createElement(SubMenuContainer, { className: "sub-container" }, item.children.map((subItem, index) => {
return (React.createElement(SubMenuItem, { onClick: () => {
this.redirect(subItem.path);
}, key: index, className: (props.location.pathname == subItem.path) ? activeClass + " sub-item" : "sub-item" },
React.createElement("span", { className: "item-label noselect" }, subItem.displayName)));
}))));
})),
!isMobile_1.default &&
React.createElement("div", { className: "top-menu" },
React.createElement(Menu, { className: "subway-menu", pose: isLarge ? 'enlarge' : 'minimize' }, props.sitemap.map((item, index) => {
let upperActiveClass = props.location.pathname == item.path ? activeClass + " item" : "item";
let upperSeenClass = this.isPredecessor(item) ? " seen" : "";
let upperClasses = upperActiveClass + upperSeenClass;
let submenuState = ((props.location.pathname == item.path) || !_.isUndefined(_.find(item.children, (i) => { return props.location.pathname == i.path; })));
return (React.createElement(React.Fragment, { key: index },
React.createElement(MenuItem, { onClick: () => {
this.redirect(item.path);
this.setState({ openSubMenu: true });
}, className: upperClasses },
React.createElement("span", { className: "item-ball" }),
React.createElement("span", { className: "item-label noselect" }, item.displayName)),
((props.location.pathname != item.path) || !item.children) && index !== props.sitemap.length - 1 && _.isUndefined(_.find(item.children, (i) => { return props.location.pathname == i.path; })) &&
React.createElement("div", { className: "stripe" + (this.isPredecessor(item) ? " seen" : "") }),
item.children && item.children.length &&
React.createElement(React.Fragment, null,
React.createElement("div", { className: (submenuState ? "open" : "close") + " stripe post-ball" + (this.isPredecessor(item) ? " seen" : "") }),
React.createElement(SubMenuContainer, { className: "sub-container " + (submenuState ? "open" : "closed"), pose: submenuState ? "openSubMenu" : "closedSubMenu" },
React.createElement("div", { className: "rotated rotatedown progress" },
React.createElement("div", null)),
React.createElement("div", { className: "rotated rotatedown" + (this.isPredecessor(item) ? " seen" : ""), style: { marginRight: "24px", } },
React.createElement("div", null)),
item.children.map((subItem, subIndex) => {
let firstClass = index == 0 ? "first" : "";
let lastClass = index !== item.children.length - 1 ? "" : "last";
let active = (props.location.pathname == subItem.path) ? activeClass : "";
let subSeenClass = this.isPredecessor(subItem) ? " seen" : "";
let classes = `sub-item ${firstClass} ${lastClass} ${active} ${subSeenClass}`;
return (React.createElement(React.Fragment, { key: subIndex },
React.createElement(SubMenuItem, { pose: props.location.pathname == subItem.path ? "expanded" : "normal", onClick: () => {
this.redirect(subItem.path);
}, key: index, className: classes },
React.createElement(SubMenuItemBall, { className: "item-ball" },
React.createElement("span", { className: "item-label noselect" }, subItem.displayName))),
subIndex !== item.children.length - 1 &&
React.createElement("div", { className: "stripe" + (this.isPredecessor(subItem) ? " seen" : ""), style: {
width: STRIPE_WIDTH,
} })));
}),
index !== props.sitemap.length - 1 &&
React.createElement("div", { className: "rotated rotateup", style: { marginRight: "20px", } },
React.createElement("div", null))),
index !== props.sitemap.length - 1 &&
React.createElement("div", { className: (submenuState ? "open" : "close") + " stripe post-ball" }))));
})))));
}
} exports.SubwayMenu = react_router_1.withRouter(MenuComponent); //# sourceMappingURL=SubwayMenu.js.map } // default/src/isMobile.js $fsx.f[58] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); exports.default = window.outerWidth < 1027; //# sourceMappingURL=isMobile.js.map } // default/src/sitemap.js $fsx.f[209] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const _ = $fsx.r(44); const Home_1 = $fsx.r(106); const Description_1 = $fsx.r(108); const Results_1 = $fsx.r(111); const Attributions_1 = $fsx.r(131); const PublicEngagement_1 = $fsx.r(171); const partners_1 = $fsx.r(173); const Parts_1 = $fsx.r(175); const Design_1 = $fsx.r(177); const Demonstrate_1 = $fsx.r(179); const Model_1 = $fsx.r(181); const Protocols_1 = $fsx.r(183); const Safety_1 = $fsx.r(185); const Notebook_1 = $fsx.r(187); const IHP_1 = $fsx.r(188); const Future_Teams_1 = $fsx.r(192); const Achievements_1 = $fsx.r(194); const Collaborations_1 = $fsx.r(195); const TeamPage_1 = $fsx.r(199); const ProjectOverview_1 = $fsx.r(201); const LabProcessOverview_1 = $fsx.r(202); const HumanPracticesOverview_1 = $fsx.r(204); const PeopleInvolvedOverview_1 = $fsx.r(205); const Introduction_1 = $fsx.r(207); const FinalWords_1 = $fsx.r(208); let items = [
{
path: "/",
comp: Home_1.Home,
displayName: "Home"
},
{
path: "/Introduction",
comp: Introduction_1.Introduction,
displayName: "Introduction",
children: [
{
path: "/Description",
comp: Description_1.Description,
displayName: "Description & Inspiration"
},
{
path: "/Achievements",
comp: Achievements_1.Achievements,
displayName: "Achievements"
}
]
},
{
path: "/Project",
comp: ProjectOverview_1.ProjectOverview,
displayName: "Project ",
children: [
{
path: "/Design",
comp: Design_1.Design,
displayName: "Background"
},
{
path: "/Results",
comp: Results_1.Results,
displayName: "Results"
},
{
path: "/Model",
comp: Model_1.Model,
displayName: "Conjugation Model"
},
{
path: "/Demonstrate",
comp: Demonstrate_1.Demonstrate,
displayName: "Demonstration"
},
]
},
{
path: "/Lab_Process_Overview",
comp: LabProcessOverview_1.LabProcessOverview,
displayName: "Parts & Procedures",
children: [
{
path: "/Safety",
comp: Safety_1.Safety,
displayName: "Safety"
},
{
path: "/Parts",
comp: Parts_1.Parts,
displayName: "Parts Catalogue"
},
{
path: "/Experiments",
comp: Protocols_1.Protocols,
displayName: "Protocols & SOPs"
},
{
path: "/Notebook",
comp: Notebook_1.Notebook,
displayName: "Notebook"
}
]
},
{
path: "/Practices_Overview",
comp: HumanPracticesOverview_1.HumanPracticesOverview,
displayName: "Practices",
children: [
{
path: "/Human_Practices",
comp: IHP_1.IHP,
displayName: "Human Practices"
},
{
path: "/Public_Engagement",
comp: PublicEngagement_1.PublicEngagement,
displayName: "Education & Public Engagement"
}
]
},
{
path: "/Future_Teams",
comp: Future_Teams_1.FutureTeams,
displayName: "To Future iGEM Teams"
},
{
path: "/People_Involved_Overview",
comp: PeopleInvolvedOverview_1.PeopleInvolvedOverview,
displayName: "People Involved",
children: [
{
path: "/Team",
comp: TeamPage_1.TeamPage,
displayName: "Team"
},
{
path: "/Attributions",
comp: Attributions_1.Attributions,
displayName: "Attributions"
},
{
path: "/Collaborations",
comp: Collaborations_1.Collaborations,
displayName: "Collaborations"
},
{
path: "/Sponsors",
comp: partners_1.PartnersSponsors,
displayName: "Sponsors & Partners"
}
]
},
{
path: "/Final_Words",
comp: FinalWords_1.FinalWords,
displayName: "Final Words"
},
]; exports.default = items; exports.FlatSitemap = _.chain(items).map((station, stationIndex) => {
if (!station.children)
return station;
else {
if (stationIndex !== items.length - 1) {
station.nextChapter = items[stationIndex + 1];
}
let adoptedKids = _.map(station.children, (kid) => {
if (stationIndex !== items.length - 1) {
kid.nextChapter = items[stationIndex + 1];
}
return kid;
});
return _.concat(station, adoptedKids);
}
}).flatten().value(); //# sourceMappingURL=sitemap.js.map } // default/src/components/Content/home/Home.jsx $fsx.f[106] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const $ = $fsx.r(102); const gsap_1 = $fsx.r(91); const isMobile_1 = $fsx.r(58); const SVG1 = () => {
return (React.createElement("svg", { version: "1.1", id: "svg-left", xmlns: "http://www.w3.org/2000/svg", x: "0px", y: "0px", viewBox: "0 0 600 400" },
React.createElement("g", null,
React.createElement("path", { className: "blob", d: "M220.262,366.814c41.228-14.367,64.978-58.826,96.198-136.802\n\t\tc43.518-108.692,53.929-137.426,67.672-149.92s154.708-58.065,177.821-65.59C576.392,9.802,591.841,5.391,596.66-2H-2v334.452\n\t\tc16.689,8.319,35.468,14.508,56.726,18.745C98.453,359.914,179.034,381.181,220.262,366.814z" }))));
}; const SVG2 = () => {
return (React.createElement("svg", { version: "1.1", id: "svg-right", xmlns: "http://www.w3.org/2000/svg", x: "0px", y: "0px", viewBox: "0 0 600 400", style: {} },
React.createElement("g", null,
React.createElement("path", { className: "blob2", d: "M361.076,143.985c9.307,26.708,38.108,42.094,88.622,62.319\n\t\tc70.412,28.192,89.027,34.936,97.12,43.839c8.093,8.903,37.615,100.223,42.49,115.196c3.045,9.354,5.902,19.361,10.691,22.483V0\n\t\tH383.337c-5.389,10.811-9.398,22.976-12.143,36.748C365.547,65.075,351.769,117.277,361.076,143.985z" }))));
}; window.HOME_ANIMATED = false; class Home extends React.Component {
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(105);
});
}
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
this.logo = React.createRef();
this.logoText = React.createRef();
this.nextBtn = React.createRef();
this.mainText = React.createRef();
this.logoContainer = React.createRef();
window.document.body.classList.add("home-page");
}
componentWillUnmount() {
window.document.body.classList.remove("home-page");
}
animate() {
if ($("#loader").length) {
setTimeout(this.animate.bind(this), 200);
return;
}
let logoRect = this.logo.current.getBoundingClientRect();
let logoTextRect = this.logoText.current.getBoundingClientRect();
let logoContainer = this.logoContainer.current.getBoundingClientRect();
const timelineOne = new gsap_1.TimelineMax({ paused: true, repeat: true });
const timelineTwo = new gsap_1.TimelineMax({ paused: true });
timelineOne.to(this.logo.current, 150, { opacity: 1, rotation: "360", ease: gsap_1.Linear.easeNone, repeat: -1 });
let scaling = isMobile_1.default ? 0.6 : 0.3;
timelineTwo
.to(this.logoContainer.current, 1, { top: isMobile_1.default ? (window.innerHeight / 6 - logoContainer.height / 2) : (window.innerHeight / 12 - logoContainer.height / 2), scaleX: scaling, scaleY: scaling, ease: gsap_1.Linear.easeIn, repeat: 0 })
.to(this.logoText.current, 1, { opacity: 1, ease: gsap_1.Linear.easeIn, repeat: 0 })
.to(this.mainText.current, 1, { opacity: 1, scaleX: isMobile_1.default ? 3.5 : 2, scaleY: isMobile_1.default ? 3.5 : 2, ease: gsap_1.Linear.easeOut })
.to(this.nextBtn.current, 1.2, { opacity: 1, delay: 1, ease: gsap_1.Linear.easeIn });
requestAnimationFrame(() => {
timelineOne.play();
timelineTwo.play();
});
}
componentDidMount() {
if (window.HOME_ANIMATED) {
return;
}
this.animate();
}
show() {
$(".next-button").addClass("btn-visible");
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "home" },
React.createElement("div", { className: "home-content" },
React.createElement("div", { ref: this.logoContainer, className: "logo-container" },
React.createElement("img", { ref: this.logo, className: "logo no-select", src: "
" }),
React.createElement("img", { ref: this.logoText, className: "text no-select", style: { opacity: 0 }, src: "
" })),
React.createElement("div", { ref: this.mainText, id: "center-text-home", className: "no-select" },
"Sensitizing Bacteria to Antibiotics ",
React.createElement("br", null),
"by Bacterial Delivery of CRISPR/Cas")),
React.createElement(SVG1, null),
React.createElement(SVG2, null),
React.createElement("div", { ref: this.nextBtn, className: "next-btn-container" },
"\t",
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation })))));
}
} exports.Home = Home; //# sourceMappingURL=Home.js.map } // default/src/components/widgets/Buttons.jsx $fsx.f[104] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const react_router_dom_1 = $fsx.r(29); const gsap_1 = $fsx.r(91); const react_blob_1 = $fsx.r(101); const $ = $fsx.r(102); exports.NextStationButton = react_router_dom_1.withRouter(class NextStationButton extends React.Component {
constructor(props) {
super(props);
this.nextStation = () => {
this.hover(false);
const timeline = new gsap_1.TimelineMax({ paused: true });
let $thisBtn = $(".next-button > div");
$thisBtn.empty();
let thisBtn = $thisBtn[0];
thisBtn.className = "next-button";
let el = thisBtn.getBoundingClientRect();
let homePage = window.document.querySelector("body.home-page");
if (homePage) {
timeline
.to(thisBtn, 0.4, {
y: (-el.top) - el.height / 2,
x: (-el.left) - el.width / 2,
scale: 3,
ease: gsap_1.Power1.easeIn,
});
}
else {
timeline
.to(thisBtn, 0.4, {
y: (-el.top + window.innerHeight / 2) - el.height / 2,
x: (-el.left + window.innerWidth / 2) - el.width / 2,
scale: 3,
ease: gsap_1.Power1.easeIn,
});
}
timeline.to(thisBtn, 0.4, {
scale: 100,
ease: gsap_1.Power1.easeIn,
borderRadius: 0,
backgroundColor: "white",
})
.call(() => {
document.body.classList.add("rodal-open");
this.props.history.push(this.props.nextStation.path);
});
requestAnimationFrame(() => timeline.play());
};
this.state = {
next: props.nextStation
};
this.button = React.createRef();
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(103);
});
}
hover(flag) {
if (flag) {
this.button.current.classList.add("btn-hover");
}
else {
this.button.current.classList.remove("btn-hover");
}
}
render() {
return (React.createElement("div", { ref: this.button, className: "next-button noselect" },
React.createElement(react_blob_1.Blob, { onClick: this.nextStation, onMouseEnter: this.hover.bind(this, true), onMouseLeave: this.hover.bind(this, false) }, "Next")));
}
}); exports.NextChapterButton = react_router_dom_1.withRouter(class NextChapterButton extends React.Component {
constructor(props) {
super(props);
this.nextChapter = () => {
this.props.history.push(this.props.nextChapter.path);
};
this.state = {
next: props.nextStation
};
}
render() {
return (React.createElement("button", { className: "text-red-900", onClick: this.nextChapter }, "NEXT CHAPTER"));
}
}); //# sourceMappingURL=Buttons.js.map } // default/src/components/Content/Project_Description/Description.jsx $fsx.f[108] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); class Description extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(107);
});
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "Project & Inspiration References",
content: `
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4520913/" target="_blank" class="references">Tan, S. Y. and Tatsumura, Y. (2015). Alexander Fleming (1881-1950): Discoverer of penicillin . Singapore Med J. 2015; 56(7): 366-367.</a>
- <a href="http://www.cdc.gov/drugresistance/threat-report-2013" target="_blank" class="references">Centers for Disease Control and Prevention, Office of Infectious Disease Antibiotic resistance threats in the United States. (2013). Biggest threats and Data. Accessed October 1, 2019.</a>
- <a href="https://www.who.int/antimicrobial-resistance/interagency-coordination-group/IACG_final_report_EN.pdf?ua=1" target="_blank" class="references">World Health Organization (WHO) Interagency Coordination Group on Antimicrobial Resistance (IACG) (2019). No time to wait: Securing the future from drug-resistant infections. Report to the secretary-general of the United Nations.</a>
- <a href="https://doi.org/10.1016/j.ijmm.2010.04.003" target="_blank" class="references">Hulscher, M. E. J. L. et al. (2010). Antibiotic use: How to improve it? Int. J. of Med. Microbiology, Vol. 300, Issue 6, 351-356, August, 2010.</a>
- <a href="https://www.nature.com/articles/s41564-018-0327-z" target="_blank" class="references">Peters, J. M. et al. (2019). Enabling genetic analysis of diverse bacteria with Mobile-CRISPRi. Nature Microbiology 4, 244-250. .</a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "description" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Description & Inspiration"),
React.createElement("div", { className: "under-rubrik" }, "Aspire to inspire"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, " We have been working with our project for more than nine months, and we are super excited to let you know about our findings. Learn more about our project and how we got inspired to create Conjugaid here."))),
React.createElement("div", { className: "row" },
React.createElement("p", { id: "desc-quote" }, "\u201CYou wanna fight?\u201D"),
React.createElement("p", null, "This is what we ask bacteria when we expose them to antibiotics. Would you not defend yourself if asked the same question? Fighting back is something bacteria have always done, by naturally defending themselves against antibiotics released by other microorganisms. "),
React.createElement("img", { className: "mx-auto", src: "https://static.igem.org/mediawiki/2019/9/94/T--SDU-Denmark--ABresistenceloop.gif?fbclid=IwAR3CIIBDhOekiqpRoJQwQDzbnf2JwDQAwOptz4WffESQF9oLgX2XYEEXei8" }),
React.createElement("p", null,
"Ever since Alexander Fleming\u2019s discovery of penicillin, antibiotics have been the weapon of choice against pathogenic bacteria ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4520913/", target: "_blank" }, "Tan, S. Y. and Tatsumura, Y. (2015). Alexander Fleming (1881-1950): Discoverer of penicillin. Singapore Med J. 2015; 56(7): 366-367. "))),
". Since then, mankind used technologic cleverness to fight back by developing new antibiotics. Nevertheless, bacteria continue to develop resistance and potentially cause untreatable infections ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "http://www.cdc.gov/drugresistance/threat-report-2013", target: "_blank" }, "Centers for Disease Control and Prevention, Office of Infectious Disease Antibiotic resistance threats in the United States. (2013). Biggest threats and Data. Accessed October 1, 2019."))),
". "),
React.createElement("p", null,
"The current development of antibiotics cannot keep up with that of antibiotic resistance ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.nature.com/articles/431899a", target: "_blank" }, "Nathan, C. (2004). Antibiotics at the crossroads. Nature, 431, 899-902. "))),
". Consequently, the World Health Organization (WHO) has estimated that in 2050, antimicrobial resistant infections could cause 10 million deaths yearly if action is not taken."),
React.createElement("img", { className: "my-10", src: "
" }),
React.createElement("p", null,
"Furthermore, the economic burden this will have on society could resemble that of the 2008-2009 financial crisis ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.who.int/antimicrobial-resistance/interagency-coordination-group/final-report/en/", target: "_blank" }, "World Health Organization (WHO) Interagency Coordination Group on Antimicrobial Resistance (IACG) (2019). No time to wait: Securing the future from drug-resistant infections. Report to the secretary-general of the United Nations. "))),
". Thus, it is crucial that we develop novel strategies to combat antibiotic resistance."),
React.createElement("p", null,
"The 2019 paper \u201CEnabling genetic analysis of diverse bacteria with Mobile-CRISPRi\u201D by Peters et al. ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.nature.com/articles/s41564-018-0327-z", target: "_blank" }, "Peters, J. M. et al. (2019). Enabling genetic analysis of diverse bacteria with Mobile-CRISPRi. Nature Microbiology 4, 244-250. "))),
" initially inspired us to realize the potential of CRISPR in developing novel therapeutic strategies to combat antibiotic resistance. With this in mind, we were drawn to \u201Cturning back time\u201D to a world in which antibiotic resistance is no longer an issue. This thought played a vital role in paving the way for defining Conjugaid as a project. We view antibiotic resistance as a problem addressable from multiple aspects, for which reason our solution includes both therapeutic and educational approaches. "),
React.createElement("p", null,
"Therapeutically, we aimed at engineering a CRISPR/Cas-system capable of sensitizing resistant E. coli to antibiotics. This system should be delivered to the target bacteria through conjugation from E. coli vehicles. Throughout our project, we have successfully constructed a CRISPR/dCas9-system that specifically targets the gfp gene (encoding green fluorescent protein, GFP) in ",
React.createElement("i", null, "E. coli."),
" Furthermore, we were able to combine our CRISPR/dCas9-system with a conjugation based delivery system and successfully deliver this to ",
React.createElement("i", null, "E. coli."),
" Following dialogue with CRISPR-company SNIPR Biome, we discovered that utilizing CRISPR/Cas9 would be far more efficient at knocking out gfp expression. Thus, we constructed a CRISPR/Cas9-system targeting the same gene as the dCas9 variant, and observed a significant improvement in its ability to knock out gfp expression."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", null,
"We also tackled antibiotic resistance by increasing public information regarding its implications and the potential of synthetic biology to help alleviate it. Besides other public engagement activities, our main focus has been to give high school students access to relevant quality education through a lesson-plan consisting of an e-book, teaching manuals and laboratory protocols. By focusing on therapeutics as well as public information, we may not only develop novel technologies, but also educate the scientists of tomorrow. This approach creates potential to solve antibiotic resistance in an efficient manner ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://doi.org/10.1016/j.ijmm.2010.04.003", target: "_blank" }, "Hulscher, M. E. J. L. et al. (2010). Antibiotic use: How to improve it? Int. J. of Med. Microbiology, Vol. 300, Issue 6, 351-356, August, 2010. "))),
". "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", null, "Click \u2018Next\u2019 to read more about our achievements within the iGEM Competition.\t"))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Description = Description; //# sourceMappingURL=Description.js.map } // default/src/components/Content/Project/But_Does_It_work/Results.jsx $fsx.f[111] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const WavesWidget_1 = $fsx.r(110); const topWaves = [
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,96L48,128C96,160,192,224,288,240C384,256,480,224,576,213.3C672,203,768,213,864,229.3C960,245,1056,267,1152,277.3C1248,288,1344,288,1392,288L1440,288L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
}
]; class Results extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "results" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Results"),
React.createElement("div", { className: "under-rubrik" }, "Clearly and Objectively"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" },
" After getting to know the fundamental principles included in our project, you may be curious to see how we have implemented them practically. Initially, we have worked on two aspects of our project separately before combining them in the end; conjugation and CRISPR/Cas, respectively. Likewise, this section is structured in the following parts:",
React.createElement("ul", null,
React.createElement("li", null, "Results from conjugation-experiments"),
React.createElement("li", null, "Results from dCas9-experiments"),
React.createElement("li", null, "Results from experiments combining conjugation and CRISPR/dCas9"),
React.createElement("li", null, "Results from Cas9-experiments"))),
React.createElement("p", null, "Keep reading to see our results of our work.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: 1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h2", { className: "darker-border" }, "Conjugation"),
React.createElement("h3", null, "Optimization of Conjugative Transfer"),
React.createElement("p", null,
"When constructing our system, we had to keep in mind the major challenges present when establishing the presence of our CRISPR/(d)Cas9-system in colonizing bacteria of the human gut microbiome. One such challenge is to ensure the ability of ingested bacteria to sensitize an adequate proportion of bacteria to antibiotics by conjugation. Therefore, we optimized the conjugative transfer of our CRISPR/(d)Cas9-system by testing the efficiency of different conjugative plasmids. We tested three different conjugative plasmids; two R-plasmids (RP4-8, Addgene no. 79813, and pRK24, Addgene no. 51950), and one F\u2019-plasmid (from an XL1-Blue strain). As mentioned in the Background section, different conjugative plasmids may contain different oriT sequences. For our conjugation systems to work, the oriT sequences found in the non-conjugative plasmid has to be compatible with the oriT sequence found in the conjugative plasmid. This necessity was met by using the R-plasmids compatible with oriT-R (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_J01003", target: "_blank" }, "BBa_J01003"),
", designed by Golden Bear 2005) and the F-plasmid was compatible with oriT-F (",
React.createElement("a", { target: "_blank", href: "http://parts.igem.org/Part:BBa_I714030" }, "BBa_I714030"),
", designed by iGEM Team Peking 2007). As illustrated in Figure 1, three different donor bacteria were designed to contain a non-conjugative and a conjugative plasmid each. The recipient bacteria were designed to contain a low-copy number plasmid (",
React.createElement("a", { href: "http://parts.igem.org/Part:pSB4C5", target: "_blank" }, "pSB4C5"),
", designed by iGEM Team Bangalore 2006). "),
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("div", { className: "figure_container" },
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 1: Graphical overview of our three donor bacteria and recipient bacterium."),
React.createElement("p", { className: "figure_text" }, "Light blue: Donor bacterium containing a non-conjugative plasmid with oriT-F and an F\u2019-plasmid. Blue: Donor bacterium containing a non-conjugative plasmid with oriT-R and a pRK24 plasmid. Dark blue: Donor bacterium containing a non-conjugative plasmid with oriT-R and a RP4-8 plasmid. Red: Recipient bacterium carrying a resistance-conferring plasmid with RFP.")),
React.createElement("p", null, "The donor and recipient bacteria were grown to late exponential phase and mixed in a 1:1 ratio. The conjugating samples were incubated at 37\u00B0C and plated on agar plates containing appropriate antibiotics, selecting for desired transconjugants. The different transconjugants were recipients (Rs) having received either the conjugative plasmid (TCs), the non-conjugative plasmid (TNs), or both (TCNs). We sought to identify the conjugative plasmid with the highest transfer frequency of both the conjugative and the non-conjugative plasmid. In order to ensure that the presence of the conjugative plasmid is necessary for the transfer of DNA to occur, we tested transfer of the non-conjugative plasmid in the absence of RP4-8. We found that no transconjugants were formed, suggesting that the Rs did not receive the non-conjugative plasmid, and that the presence of the RP4-8 plasmid was necessary for transfer of the non-conjugative plasmid. We also constructed bacteria containing a non-conjugative plasmid lacking the oriT site, along with the RP4-8 plasmid, in order to confirm that presence of oriT is required for transfer of the non-conjugative plasmid. Similarly, we found that no Rs had received the non-conjugative plasmid, confirming that oriT is required. "),
React.createElement("p", null,
"Based on the data shown in ",
React.createElement("span", { className: "font-bold" }, "Figure 2"),
", a relatively uniform dispersal of either the non-conjugative plasmid (TN), the conjugative plasmid (TC) or both plasmids (TCN) are obtained when utilizing the RP4-8 plasmid. The pRK24 plasmid (Addgene, no. 51950) seems to favor the delivery of the non-conjugative plasmid rather than its own transfer to recipient bacteria, whereas the F\u2019-plasmid seems to favor its own transfer rather than facilitating the transmission of the non-conjugative plasmid. The highest proportion of transconjugants having received both the non-conjugative and the conjugative plasmid is observed for the experiments made with the RP4-8 plasmid. However, no significant difference in the formation of either TCs, TNs, or TCNs is observed for the experiments based on the two-way ANOVA with Tukey\u2019s multiple comparisons test."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 2: The proportion of cells having received either the conjugative plasmid (TCs), cells having received the non-conjugative donor plasmid (TNs) or cells having received both plasmids (TCNs)."),
React.createElement("p", { className: "figure_text" }, "The proportion of TCs, TNs and TCNs were obtained by dividing the number of CFUs with the total number of recipient cells. Both donor and recipient cells are E. coli K12 TOP10. Donor cells contain the pRK24, RP4-8 or F\u2019 conjugative plasmid along with a non-conjugative plasmid containing a compatible oriT sequence, and recipient cells contain a low-copy number plasmid. Data are represented as mean values with standard error of mean (SEM). A two-way ANOVA with Tukey\u2019s multiple comparisons test revealed no significant differences between groups (N=4-8).")),
React.createElement("p", null,
"The RP4-8 plasmid has a broad host range indicating that the conjugative system can be delivered to a diverse population of bacterial strains ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Schaeter, M. (2009) Desk Encyclopedia of Microbiology. Oxford & San Diego: Elsevier. Pages 296-297."))),
". As the intestines harbour a large diversity of bacterial strains ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1395357/", target: "_blank" }, "Eckburg, P. B. et al. (2006) Diversity of the Human Intestinal Microbial Flora. Science, 308(5728): 1635-1638. "))),
", this feature serves as an advantage when evaluating the plasmid in a clinical context. Aside from the broad host range, the mating efficiency is considered high due to the constitutively active system of the RP4-8 plasmid ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Schaeter, M. (2009) Desk Encyclopedia of Microbiology. Oxford & San Diego: Elsevier. Pages 296-297."))),
". When considering these known features of the RP4-8 plasmid, including the data obtained in the experiments on the conjugative plasmid in combination with the non-conjugative plasmid, we decided to base future experiments on the RP4-8 plasmid."),
React.createElement("h3", null, "Dispersal of Plasmid Over Time"),
React.createElement("p", null,
"Clinically, it is of relevance to ensure a sufficient dispersal of our CRISPR/(d)Cas9-system to resistant intestinal bacteria within a limited timeframe. This is especially crucial when considering patients already diseased by infections with antibiotic resistant bacteria. In that regard, we wanted to examine the transmission of the non-conjugative plasmid containing the oriT-R sequence over time. We chose to exclude the CRISPR/(d)Cas9-system from the non-conjugative plasmid, assuming that this does not alter the plasmid transfer rate. The experiments were carried out in a bacterial mixture that, albeit simplified, resembles the conditions of the natural human gut microbiome. This includes cultivation of bacteria in liquid solution and high cell density. However, as literature on conjugation protocols for interrupted mating at different time points in liquid solution is scarce, we sought to develop a ",
React.createElement("a", { href: "https://static.igem.org/mediawiki/2019/c/c0/T--SDU-Denmark--SOP8.pdf", target: "_blank" }, "procedure"),
" allowing us to examine how plasmids spread over time. To follow the progress on the optimization of this protocol, have a look at the ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Notebook", target: "_blank" }, "Notebook for Conjugation"),
". "),
React.createElement("p", null,
"The donor bacteria chosen from the previous experiment containing the RP4-8 plasmid and the non-conjugative plasmid with the oriT-R sequence, as well as the recipient bacteria, were used for examining conjugation frequency over time. The donor and recipient bacteria were grown to late exponential phase and mixed in a 1:1 ratio. The conjugating sample was allocated into separate tubes and incubated at 37\u00B0C under rotation. At two hour intervals, the samples were plated on agar plates containing appropriate antibiotics, selecting for desired transconjugants. The outcome of this experiment is seen in",
React.createElement("span", { className: "font-bold" }, " Figure 3"),
". "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 3: The proportion of cells having received either the conjugative plasmid (TCs), cells having received the non-conjugative donor plasmid (TNs) or cells having received both plasmids (TCNs)."),
React.createElement("p", { className: "figure_text" }, "CFU was normalized to the total number of recipients as a function of time. The conjugative plasmid used for this data collection is RP4-8 that is compatible to the oriT sequence found within the non-conjugative plasmid.")),
React.createElement("p", null,
"The formation of the different transconjugants increases somewhat linearly with time as observed in ",
React.createElement("span", { className: "font-bold" }, "Figure 3"),
". However, the system seems to slightly favor the transfer of either the conjugative plasmid or the non-conjugative plasmid. Approximately 3.5% of total recipients received either plasmid after 10 hours, rather than both plasmids simultaneously (~3%), reflected by the formation of TCs, TNs and TCNs, respectively. The formation of the transconjugants is affected by the ability of TNs to remain susceptible towards the conjugative plasmid, thus becoming TCNs. However, the TCs do not share this ability as they are unable to function as recipients for the non-conjugative plasmid, due to a phenomenon known as fertility inhibition, by which conjugation of co-resident plasmids within donor cells is prevented ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/18440635", target: "_blank" }, "Garcillan-Barcia, M. P. and F. de la Cruz (2008). Why is entry exclusion an essential feature of conjugative plasmids? Plasmid 60(1): 1-18."))),
". "),
React.createElement("p", { id: "no-bottom" },
"Although these results provide information on the rate of plasmid transfer and dispersal of the transconjugant population over time, we recognize the limitations of our system compared with the actual human gut microbiome. We are aware that the conditions of the intestines cannot be fully replicated in vitro, and that many factors may affect conjugation frequency, such as pH, salinity, O2 levels and nutrient availability ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/3319117/", target: "_blank" }, "Khalil, T. A. et al. (1987). Temperature, pH, and cations affect the ability of Escherichia coli to mobilize plasmids in L broth and synthetic wastewater. Can J Microbiol, 33(8), 733-737."))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/3312987/", target: "_blank" }, "Roszak, D. B. et al. (1987). Survival strategies of bacteria in the natural environment. Microbiol Rev, 51(3), 365-379."))),
". Thus, additional measures can be taken to further mimic the gastrointestinal tract, such as using continuous flow fermentors and multistage chemostats ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/10721913", target: "_blank" }, "Gibson, G. R. et al. (2000). Aspects of in vitro and in vivo research approaches directed toward identifying probiotics and prebiotics for human use. J Nutr, 130(2S Suppl), 391S-395S."))),
", which would be relevant to include in future iterations of our system. Nonetheless, when taking into account the available resources and timeframe of our project, our system provides valuable insight into the dispersal of conjugative and non-conjugative plasmids in the human intestine. In a clinical context, however, it is relevant to acquire information on the dispersal of our conjugative and non-conjugative plasmids over even longer periods of time, in order to obtain knowledge of the time required to sensitize a sufficient proportion of the gut bacteria. This issue will be further addressed in our Modelling section, where we model the kinetics of conjugative plasmid transfer. For now, keep reading to see results from our work with dCas9-experiments.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("h2", { className: "whiter-border" }, "CRISPR/Cas"),
React.createElement("h3", null, "Targeting gfp with dCas9"),
React.createElement("p", null,
"To downregulate expression of gfp we initially sought to utilize the CRISPR/dCas9-system rather than the catalytically active Cas9 variant. This decision was based on literature supporting Cas9 possibly resulting in cell death when targeting chromosomal DNA . Even though our current aim was to target antibiotic resistance genes carried on plasmids, utilizing dCas9 might prove beneficial when targeting resistance genes carried on chromosomal DNA in future iterations of our project. For that purpose, we codon optimized the ",
React.createElement("a", null, "BBa_K1150000 "),
" part from the iGEM kit for use in our chassis E. coli K12 TOP10 and assembled dCas9+sgRNA-systems targeting gfp. We constructed two different target bacteria containing gfp; one with a strong promoter and one with a weak promoter. To mimic OXA-48 plasmids, we transferred gfp to a low copy number plasmid backbone (",
React.createElement("a", null, "pSB4C5"),
"). Thus, our two target bacteria are as follows:"),
React.createElement("ol", null,
React.createElement("li", null,
"GFP with a strong constitutive Anderson promoter with a strength of 0.86 (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K1150000", target: "_blank" }, "BBa_K3044029"),
"). This target bacteria will henceforth be referred to as gfp 0.86."),
React.createElement("li", null,
"GFP with a weak constitutive Anderson promoter with a strength of 0.33 (",
React.createElement("a", { href: "http://parts.igem.org/Part:pSB4C5", target: "_blank" }, "BBa_K3044002"),
"). This target bacteria will henceforth be referred to as gfp 0.33.")),
React.createElement("p", null,
"To investigate the efficiency of gfp repression by dCas9, two different CRISPR/dCas9-systems were constructed, as illustrated in Figure 4. These systems contained two different sgRNAs, each targeting gfp in the 5\u2019-end of the DNA strand. One sgRNA (",
React.createElement("a", { target: "_blank", href: "http://parts.igem.org/Part:BBa_K3044010" }, "BBa_K3044010"),
", henceforth referred to as sgRNA1) targets the template strand, while the other sgRNA (",
React.createElement("a", { target: "_blank", href: "http://parts.igem.org/Part:BBa_K3044011" }, "BBa_K3044011"),
", henceforth referred to as sgRNA2) targets the non-template strand. Each of the two sgRNAs were inserted into our plasmid containing dCas9, forming our two dCas9+sgRNA-systems targeting gfp (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044013", target: "_blank" }, "BBa_K3044013"),
" and ",
React.createElement("a", { target: "_blank", href: "http://parts.igem.org/Part:BBa_K3044014" }, "BBa_K3044014"),
"). The dCas9+sgRNA-systems were each combined with an IPTG inducible promoter to enable control of its expression. "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 4: Two dCas9/sgRNA-systems targeting gfp"),
React.createElement("p", { className: "figure_text" }, "The dCas9/sgRNA1 system targeted the template strand and the dCas/sgRNA2 system targeted the non-template strand.")),
React.createElement("p", null,
"The plasmids containing dCas9+sgRNA1 and dCas9+sgRNA2 were transformed into gfp 0.86 and gfp 0.33. In order to verify the effect of the combined dCas9+sgRNA-systems, as opposed to the effect of the individual components, the sgRNAs (sgRNA1 and sgRNA2) as well as dCas9 (",
React.createElement("a", null, "BBa_K3044009"),
") were transformed into our target bacteria. To evaluate the efficiency of our dCas9+sgRNA-systems in downregulating the expression of gfp, fluorescence microscopy was performed in combination with fluorescence-activated cell sorting (FACS). The transformants were grown over night with IPTG to fully induce dCas9 expression. As LB medium can autofluoresce, samples from the overnight culture (ON) were washed in PBS buffer before measuring by FACS. Based on the FACS analysis, a histogram was plotted as illustrated in Figure 5 illustrating the forward scatter area for each cell in order to examine the distribution of the bacterial populations. Cells with a forward scatter value below zero were excluded from the plot."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 5: Cell count as a function of Forward Scatter."),
React.createElement("p", { className: "figure_text" }, "Cell size plot for eight different bacteria populations (E.coli WT, gfp 0.86, gfp 0.86+dCas9, gfp 0.86+Cas9+sgRNA1, gfp 0.33, gfp 0.33+dCas9+sgRNA1, gfp 0.33+dCas9, and gfp 0.33+Cas9+sgRNA1) showing the normal distribution of the bacterial populations.")),
React.createElement("p", null, "Based on the FACS data shown in Figure 5, it seems that all populations follow a Gaussian distribution. For this reason, we allow ourselves to assume that the cell populations also follow a Gaussian distribution when evaluated in regard to fluorescence. Based on this plot, the mean was set to 180 (arbitrary) with a standard deviation of 100 for all populations."),
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("div", { className: "figure_container" },
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 6:"),
React.createElement("p", { className: "figure_text" }, "A: Microscopy image of the different (d)Cas9-systems and a positive and negative control transformed into the target bacteria gfp 0.86. Upper row: Phase contrast microscopy images (20 ms exposure time). Middle row: Fluorescence microscopy images (50 ms exposure time). Lower row: Merged images of phase contrast and fluorescence. The different columns represent the WT, the target bacterium (gfp 0.86) and the systems transformed into gfp 0.86. This includes the different sgRNAs (sgRNA1 and sgRNA2), the two dCas9/sgRNA-systems (dCas9+sgRNA1 and dCas9+sgRNA2), dCas9+oriT, dCas9+sgRNA+oriT as well as Cas9+sgRNA1. B: Histograms of GFP/area signal vs. cell count of the mean of the three biological replicates generated from the FACS. The histograms compare the E. coli WT (grey) and the target bacterium gfp 0.86 (green) with either dCas9, sgRNA1, sgRNA2, the two (d)Cas9/sgRNA-system and the Cas9/sgRNA-system (all in blue). C: Barchart showing average GFP absorbance per cell area normalized to gfp 0.86 control. Data are represented as mean values with standard deviation (sd). A one-way ANOVA with Dunnett\u2019s multiple comparisons test revealed a highly significant difference between the gfp 0.86 control and the WT (p0.001). Cas9 + sgRNA1 was able to lower GFP absorbance to the level of the WT (p<0.001), whereas the remaining constructs showed no significant difference compared with the gfp 0.86 control (p<0.05).")),
React.createElement("img", { id: "", className: "", src: "https://static.igem.org/mediawiki/2019/7/73/T--SDU-Denmark--AFSF033.png?fbclid=IwAR0GOP2PCqxyYSoHQhIbxqr-ZPFu2bwlOTyqe35O92nYEK3zFN4veiIpeCQ" }),
React.createElement("div", { className: "figure_container" },
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 7: "),
React.createElement("p", { className: "figure_text" }, "A: Microscopy image of the different (d)Cas9-systems and a positive and negative controls transformed into the target bacteria gfp 0.33. Upper row: phase contrast microscopy images (20 ms exposure time). Middle row: fluorescence microscopy images (50 ms exposure time). Lower row: merged images of phase contrast and fluorescence. The different columns represent the WT, the target bacterium (gfp 0.33) and the systems transformed into gfp 0.33. This includes the different sgRNAs (sgRNA1 and sgRNA2), the two dCas9/sgRNA systems (dCas9+sgRNA1 and dCas9+sgRNA2), dCas9+oriT, dCas9+sgRNA+oriT as well as Cas9+sgRNA1. B: Histograms of GFP/area signal vs cell count of the mean of the three biological replicates FACS. The histograms compare the E. coli WT (grey) and the target bacterium gfp 0.33 (green) with either dCas9, sgRNA1/sg or a (d)Cas9/sgRNA-system. C: Barchart showing average GFP absorbance per cell area normalized to gfp 0.33 control. Data are represented as mean values with standard deviation (sd). A one-way ANOVA with Dunnett\u2019s multiple comparisons test revealed a highly significant difference between the gfp 0.33 control and the WT (p<0.001). Cas9 + sgRNA1 was able to lower GFP absorbance to the level of the WT (p<0.001), whereas the remaining constructs showed no significant difference compared with the gfp 0.33 control (p<0.05).")),
React.createElement("p", null,
"Neither dCas9 nor sgRNA1 or sgRNA2 alone showed any effect on the level of GFP fluorescence, supported by both the fluorescence images in Figure 6A and Figure 7A as well as the FACS analysis in Figure 6B+C and Figure 7B+C. This was expected, as the ability of CRISPR/dCas9 to repress gene expression requires simultaneous presence of both the dCas9 protein and the sgRNA strand. Based on the fluorescence images shown in Figure 6A and Figure 7A, it seems that both the dCas9+sgRNA1 and dCas9+sgRNA2 constructs are able to lower the level of GFP fluorescence in both the gfp 0.86 and gfp 0.33 target bacteria. However, this observation was not supported by the FACS analysis, as neither construct was able to significantly affect the average level of GFP fluorescence in either gfp 0.86 and gfp 0.33, (p< 0.05), based on a one-way ANOVA with Dunnett\u2019s multiple comparisons test. Thus, it seems that the different expression levels of GFP in the target bacteria did not influence the efficiency of the dCas9+sgRNA constructs in repressing fluorescence. We reckoned that the ability of our dCas9+sgRNA constructs to reduce GFP fluorescence would show greater effect when GFP is highly expressed due to a high transcriptional activity causing a greater potential for downregulation, however, this was not the case. Furthermore, it seems that neither sgRNA1 nor sgRNA2 in combination with dCas9 are effective in repressing GFP fluorescence in either target bacteria. We expected both sgRNAs to provide a relatively high level of gene repression as both of these target the 5\u2019-end of the DNA strand. This would potentially block transcriptional elongation close to the transcription start site, thereby preventing any functional transcript of the GFP gene from being produced. From Figure 6 and Figure 7 it does not seem to make a difference for our system whether the sgRNA targets the template (sgRNA1) or the non-template (sgRNA2) strands. This is opposing literature indicating that a more efficient repression of gene expression is obtained when targeting the non-template strand rather than the template strand ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3922765/", target: "_blank" }, "Larson, M. H. et al. (2013). CRISPR interference (CRISPRi) for sequence-specific control of gene expression. Nat Protoc, 8(11), 2180-2196. "))),
". Therefore, neither of the dCas9 systems targeting gfp worked as expected. This was despite dCas9 being expressed on a high copy number plasmid and gfp on a low copy number plasmid, likely ensuring a sufficient level of dCas9 to repress gfp transcription. Since dCas9 requires continuous binding to the DNA strand to obtain efficient gene repression, the constant dissociation and association with the DNA, determined by the binding constant of the dCas9/sgRNA-system, could allow for some transcription of the target gene to occur. Furthermore, the lack of GFP repression could be a potential consequence of the high stability and long half-life of GFP, as even a slight remaining transcriptional activity of gfp might be sufficient for the cell to obtain a normal level of GFP expression ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/10611396", target: "_blank" }, "Corish, P. and Tyler-Smith C. (1999). Attentuation of green fluorescent protein half-life in mammalian cells. Protein Eng, 12(12), 1035-1040. "))),
". As GFP fluorescence was evaluated from overnight (ON) cultures, this could have allowed sufficient time for the cells to obtain a steady level of GFP, thus reaching similar levels as control cells. Evaluating GFP fluorescence from bacteria in exponential phase might therefore provide a way to circumvent this issue. In this phase, GFP might not have accumulated, and a more representative GFP signal might be seen. "),
React.createElement("h3", null, "Combining Conjugation with CRISPR/dCas9"),
React.createElement("p", null,
"We have proven that the dispersal of conjugative and non-conjugative plasmids increase over time and that we succeeded in constructing a CRISPR/dCas9-system optimized for expression in ",
React.createElement("i", null, "E. coli."),
" Based on this, we sought to combine the two systems. To obtain this goal, the ",
React.createElement("span", null, "dCas9+sgRNA plasmid"),
" (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044013", target: "_blank" }, "BBa_K3044013"),
") was assembled with the oriT-R part (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_J01003", target: "_blank" }, "BBa_J01003"),
") used for the former conjugation experiments. Besides the CRISPR/dCas9-oriT construct, a similar plasmid lacking the sgRNA sequence was designed as a control to prove the necessity of the sgRNA for optimal functionality of the dCas9 protein. Both plasmids were designed to contain the oriT-R part downstream of the dCas9 sequence in the plasmid backbone. To confirm correct insertion of oriT at this location, specific primers were used for the PCR reaction. The forward primer was designed to hybridize around position 2500 in the nucleotide sequence of the dCas9 gene. The reverse primer was designed to hybridize in the suffix region of the plasmid backbone. The customized primer designs were manufactured to distinguish a successful elongation of the short oriT-R sequence of 400 nucleotides relative to a 4100 nucleotide dCas9 sequence. The gel image in ",
React.createElement("span", null, "Figure 8"),
" reveals successful assembly of the two plasmids. The first lanes (1-4) show the presence of the oriT sequences in both plasmid constructs (2200), whereas the last lanes (6-9) show the presence (1000) and absence (850) of the sgRNA sequence in the two CRISPR/dCas9-oriT constructs (one lacking the sgRNA sequence), respectively."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 8: Agarose gel (1% agarose) of PCR products."),
React.createElement("p", { className: "figure_text" }, "Lane 1-4 verifies the presence of the oriT sequence downstream of the dCas9 sequence, whereas lane 1-2 examines BBa_K3044016 containing the dCas9+sgRNA+oriT construct, and lane 3-4 examines BBa_K3044028 containing the dCas9-oriT system. Lane 6-9 verifies the presence and absence of the sgRNA sequence in BBa_K3044016 and BBa_K3044028, respectively. L indicates the 10 kb DNA size marker.")),
React.createElement("p", null,
"To enable transfer of the CRISPR/dCas9-oriT construct from our donor bacteria, the RP4-8 plasmid was conjugated into the donor bacteria according to the same procedure as for the conjugation experiments covered previously (",
React.createElement("span", null, "Figure 2 and 3"),
"). The new donor bacteria act as suppliers of both the non-conjugative and the RP4-8 plasmid to the target bacteria each containing a plasmid encoding ",
React.createElement("span", null, "gfp"),
" (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044029", target: "_blank" }, "BBa_K3044029"),
"). Conjugation was carried out by growing donor and recipient cells to late exponential phase followed by mixing them in a 1:1 ratio. The conjugating samples were incubated at 37\u00B0C and plated on agar plates containing appropriate antibiotics selecting for transconjugants. In order to verify the presence of both the ",
React.createElement("span", null, "dCas9+sgRNA+oriT"),
" and the ",
React.createElement("span", null, "Cas9+oriT"),
" plasmid as well as the plasmid containing gfp in the transconjugants, a PCR reaction was performed. Three primers were used for this purpose; the two primers used for the previous PCR reaction, which multiply part of the dCas9 sequence including the oriT sequence, and a third primer hybridizing to the prefix region of the plasmid backbone, which multiplies the gfp sequence along with the primer hybridizing to the suffix region. The gel image in ",
React.createElement("span", null, "Figure 9"),
" reveals successful delivery of both the ",
React.createElement("span", null, "dCas9+sgRNA+oriT"),
" plasmid to recipient cells containing the gfp plasmid construct. The first lanes (1-8) show the presence of the ",
React.createElement("span", null, "dCas9+oriT"),
" construct (2200), transferred from the donor, as well as the plasmid containing gfp (1200) found within the recipient cell itself. The last lanes (9-12) show the presence of gfp only for recipient cells that have not received the ",
React.createElement("span", null, "dCas9+oriT"),
" plasmid (1200). "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 9: Agarose gel (1% agarose) of PCR products."),
React.createElement("p", { className: "figure_text" }, " Lane 1-4 verifies the presence of the dCas9+sgRNA+oriT plasmid received from the donor bacterium and the plasmid containing the gfp gene. Lane 5-8 verifies the presence of the dCas9+oriT plasmid received from the donor bacterium acting as a control for the action of CRISPR/dCas9. Lane 9-12 display the presence of the plasmid containing the gfp in recipient cells. L indicates the 10 kb DNA size marker. ")),
React.createElement("p", null,
"In agreement with ",
React.createElement("span", null, "Figure 9"),
" above, it can be concluded that the dispersal of the ",
React.createElement("span", null, "dCas9+sgRNA+oriT"),
" and the ",
React.createElement("span", null, "dCas9+oriT"),
" plasmids is the result of the action of the RP4-8 plasmid."),
React.createElement("h3", null, "Targeting gfp with Cas9"),
React.createElement("p", null,
"Since we did not achieve any significant reduction in GFP fluorescence when targeting gfp 0.86 and gfp 0.33 with our dCas9+sgRNA-system, we initiated discussions with CRISPR-company SNIPR Biome and chose to assemble a Cas9+sgRNA-system targeting gfp after receiving advice from them. As CRISPR/dCas9 represses the target gene instead of knocking it out, it needs to be continuously expressed in the cell to maintain a low level of gene expression. However, Cas9 induces a complete knockout of the target gene, for which reason it does not need to be permanently expressed in our target bacteria. Our Cas9+sgRNA-system was assembled by transforming our dCas9 part (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044009", target: "_blank" }, "BBa_k3044009"),
") into Cas9 by introducing two mutations using Phusion PCR. Our CRISPR/Cas9-system targeting gfp contains sgRNA1. When testing the Cas9+sgRNA1-system, experiments identical to those for dCas9 were performed. The fluorescence images in Figure 6 and Figure 7 indicates that the Cas9+sgRNA1-system is able to reduce GFP fluorescence in both gfp 0.86 and gfp 0.33 to the level of the WT which is also indicated by the FACS histograms in Figure 6 and Figure 7. The ANOVA-test shows a significant knockout of the gfp expression for gfp 0.86 with p< 0.05. "),
React.createElement("p", null, "The knockout of gfp by the Cas9+sgRNA1-system for both gfp 0.86 and gfp 0.33 is more efficient than repression by the dCas9 systems. To identify when Cas9 works, a time experiment measuring percentage fluorescence in the population was performed using FACS. This was done to examine at which phase the Cas9+sgRNA1-system was most effective. OD600 and FACS were measured with one hour intervals for 5 hours. To interpret the FACS data, it was assumed that all populations were distributed normally based on a cell size plot (similar to data in Figure 5). The data is plotted with percentage fluorescence over time and OD when comparing the wild-type (WT) and Cas9+sgRNA1-system with gfp 0.86 as illustrated in Figure 10. Fluorescence is seen for gfp 0.86 (Figure 10A) at all time intervals and OD600 values. This is expected as this strain is the positive fluorescence control. Minor fluorescence is observed for the WT (Figure 10B) at the beginning and end of the experiment. This is caused by the division of the GFP-absorbance with cell size. In the lag- and stationary phase the cell size of the bacteria is smaller, giving rise to minimal fluorescence observation, caused by uncertainty in the FACS measurements. Through the remaining time intervals and OD600 values, no fluorescence is observed as expected, as the cell size of the bacteria increases through the exponential phase. Cas9+sgRNA1 (Figure 10C) shows no fluorescence through the plot, except for in the first measurement. This is also caused by a smaller cell size in the lag phase compared to the later phases. By comparing these three plots, it is observed that knockout of gfp 0.86 gene expression takes place at the first cell division after insertion of the Cas9+sgRNA1-system causing a knockout of gfp in this strain."),
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("div", { className: "figure_container" },
React.createElement("p", { className: "figure_name text-blue-800" },
"Figure 10: Percentage fluorescent bacteria at time intervals to different OD",
React.createElement("sub", null, "600"),
" values."),
React.createElement("p", { className: "figure_text" },
"Facs data for gfp 0.86 (A), WT (B) and gfp 0.86+Cas9+sgRNA1 (C). Fluorescence is seen for gfp 0.86 (A) at all time intervals and OD",
React.createElement("sub", null, "600"),
" values. ")),
React.createElement("p", null, "From these results, it is clear that the Cas9+sgRNA1-system has a significantly higher effect regarding gfp repression, as expected, compared to the dCas9+sgRNA-systems. Since the target gene is knocked out upon Cas9 cleavage, the cell is dependent on precise cell repair mechanisms in order to express the gene. The observed decrease in fluorescence indicates that a Cas9 attack in a gene expressed from a low copy number plasmid can dramatically repress the expression of this gene. Since Cas9 has been shown to effectively knockout the expression of fluorescence genes, it can be argued that Cas9 will have the same effect on other genes, such as genes encoding antibiotic resistance."))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: 1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h2", { className: "darker-border" }, "Additional Results"),
React.createElement("h3", null, "Targeting rfp with dCas9 and Cas9"),
React.createElement("p", null,
"In addition to targeting gfp, we constructed (d)Cas9+sgRNA-systems against rfp (encoding red fluorescent protein, RFP) to test whether our systems could repress the expression of different target genes. Due to time and resource limitations, however, we focused primarily on gfp, for which reason we did not obtain the same amount of results for experiments testing rfp. For our experiments, our CRISPR/(d)Cas9-systems targeting rfp were transformed into our target bacteria expressing RFP with a strong constitutive Anderson promoter (strength 0.86, ",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_J23102", target: "_blank" }, "BBa_J23102"),
") targeting the non-template strand closest to the 3\u2019-end of the gene. Henceforth, this target bacterium will be referred to as rfp 0.86. When testing the effect of the dCas9+sgRNA-systems targeting rfp, the dCas9+sgRNA (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044021", target: "_blank" }, "BBa_K3044021"),
"), the Cas9+sgRNA-system (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044026", target: "_blank" }, "BBa_K3044026"),
"), dCas9 alone as well as the individual sgRNA were all transformed into rfp 0.86. As for the dCas9-systems targeting rfp, fluorescence microscopy was performed on the ON cultures of transformants to obtain qualitative data for the efficiency, from which images can be seen in Figure 11."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 11: Fluorescence microscopy images of the different constructs and controls transformed into rfp 0.86. :"),
React.createElement("p", { className: "figure_text" }, "Upper rowPhase contrast microscopy images (20 ms exposure time). Middle row: Fluorescence microscopy images (100 ms exposure time). Lower row: Merged images of phase contrast and fluorescence. The different columns represent the WT, the target bacteria (rfp 0.86), and the systems transformed into rfp 0.86, including the sgRNA, dCas9, dCas9/sgRNA system and the Cas9+sgRNA.")),
React.createElement("p", null, "In contrast to gfp, a decrease in fluorescence is observed in bacteria having received either the sgRNA or dCas9 protein alone. However, a more efficient repression is observed when the target bacteria receive a complete (d)Cas9+sgRNA-system, since a minimal level of fluorescence is observed in these bacteria. Due to time and resource limitations, qualitative data for the (d)Cas9-systems targeting rfp was not achievable. Therefore, the efficiency of these systems remains inconclusive. "),
React.createElement("h3", null, "Targeting Chloramphenicol Resistance with dCas9"),
React.createElement("p", null,
"In addition to targeting fluorescence genes with our dCas9 system, we constructed dCas9 systems targeting the gene encoding chloramphenicol (CML) resistance as well, as illustrated in Figure 12. We ordered sgRNAs targeting different sequences in the resistance gene and inserted them in front of dCas9 (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044009", target: "_blank" }, "BBa_K3044009"),
"). We finished four different constructs of dCas9/sgRNA-system targeting CML resistance."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 12"),
React.createElement("p", { className: "figure_text" }, "Assembly of sgRNA (targeting CML resistance) and BBa_K3044009. The primers used in the PCR hybridized to prefix and the SalI restriction site in the 5\u2019-end of dCas9. For successful ligations we expect lengths on approximately 1000 bp while the PCR fragment of the BBa_K3044009 part alone is approximately 800 bp.")),
React.createElement("p", null, "Due to time limitations we did not test these systems since we worked on other proof-of-concept experiments. After evaluating the results of our dCas9- and Cas9-systems, it became clear that the sgRNAs targeting CML resistance should have been implemented into a Cas9-system if time allowed it. Hereafter, MIC (minimum inhibitory concentration) experiments should be performed to investigate the ability of our system to knock out resistance genes in CML resistant bacteria."),
React.createElement("p", { id: "no-bottom" }, "Besides performing experiments in the laboratory, we have modelled the kinetics of plasmid transfer in conditions resembling the human gut microbiome to an extent. Click \u2018Next\u2019 to learn more.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Results = Results; //# sourceMappingURL=Results.js.map } // default/src/components/widgets/WavesWidget.jsx $fsx.f[110] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const react_1 = $fsx.r(3); const _ = $fsx.r(44); { } const WIDTH_SCALING_FACTOR = Math.pow(10, 4); const modifyWaves = (waves) => {
return _.map(waves, (svg) => {
let responsiveHeight = 1 + (1536 - window.innerHeight) / WIDTH_SCALING_FACTOR;
svg.offset = svg.offset * 1 / responsiveHeight;
return svg;
});
}; const getInterWaveOffsetFactor = (index) => {
let responsiveFactor = 1; return (index + 1) / responsiveFactor;
}; const getWaveSectionHeight = (waves, svgRefs) => {
let values = [];
let svgs = waves;
return _.chain(svgs).map((svg, index) => {
let responsiveHeight = 1 + (1536 - window.innerHeight) / WIDTH_SCALING_FACTOR;
let divHeight = svg.offset;
if (index == svgs.length - 1) {
divHeight += svgRefs[index].current.clientHeight;
}
return divHeight;
}).reduce((sum, svg) => sum + svg).value();
}; const getWaveTrueOffset = (waves, index) => {
return _.chain(waves.slice(0, index + 1)).map((wave) => {
return wave.trueOffset ? wave.trueOffset : 0;
}).reduce((sum, val) => sum + val).value();
}; const updateDimensions = (setSectionHeight, setBlockHeights, svgRefs, heightRefs, waves, topOffset) => {
let allSvgLoaded = true;
_.each(svgRefs, (svg, index) => {
if (!svg.current) {
allSvgLoaded = false;
return;
}
let height = svg.current.height.baseVal.value;
if (height <= 0) {
allSvgLoaded = false;
}
});
if (!allSvgLoaded) {
setTimeout(updateDimensions.bind(this, setSectionHeight, setBlockHeights, svgRefs, heightRefs, waves, topOffset), 50);
}
else {
let waveSectionHeight = getWaveSectionHeight(waves, svgRefs);
let sectionHeight = waveSectionHeight + topOffset;
setSectionHeight(parseInt(sectionHeight));
let isUpdated = false;
let diffs = _.map(heightRefs, (ref, index, refs) => {
let isLast = index == (refs.length - 1);
let blockSizing = ref.current.getBoundingClientRect();
let firstParent = ref.current.parentElement;
let nextElement = isLast ? firstParent.parentElement.nextElementSibling : firstParent.nextElementSibling;
let isReversed = waves[index].reversed;
if (nextElement) {
let nextElementSizing = nextElement.getBoundingClientRect();
if (isLast) {
let svgContribution = isReversed ? svgRefs[index].current.getBoundingClientRect().height : 0;
isUpdated = (nextElementSizing.top - (blockSizing.top + blockSizing.height + svgContribution)) <= 0.1;
}
let waveTopOffsetDiff = Math.abs(waves[index].offset - (index == 0 ? 0 : waves[index - 1].offset));
let diff = (isReversed && isLast) ? Math.abs(nextElementSizing.top - blockSizing.top) - svgRefs[index].current.getBoundingClientRect().height : Math.abs(nextElementSizing.top - blockSizing.top);
return diff > 0 ? diff + waveTopOffsetDiff : 0 + waveTopOffsetDiff;
}
});
setBlockHeights(diffs);
if (!isUpdated) {
setTimeout(updateDimensions.bind(this, setSectionHeight, setBlockHeights, svgRefs, heightRefs, waves, topOffset), 100);
}
}
}; exports.Waves = function (props) {
$fsx.r(109);
const [sectionHeight, setSectionHeight] = react_1.useState(0);
const [blockHeights, setBlockHeights] = react_1.useState([]);
const trueOffsets = _.map(props.waves, (wave, index, waves) => {
return getWaveTrueOffset(waves, index);
});
let svgRefs = _.map(props.waves, () => react_1.useRef());
let heightRefs = _.map(props.waves, () => react_1.useRef());
let zIndexBase = props.waves.length + (props.concurrentZDelta ? -props.concurrentZDelta : 1) * 10;
react_1.useLayoutEffect(() => {
let update = updateDimensions.bind(this, setSectionHeight, setBlockHeights, svgRefs, heightRefs, props.waves, props.top);
window.addEventListener('resize', _.throttle(update, 100));
update();
return () => {
window.removeEventListener('resize', _.throttle(update, 100));
};
}, []);
return (React.createElement("div", { style: {
height: sectionHeight,
overflow: "hidden",
position: "relative",
top: `${props.top ? props.top : 0}px`
} },
props.children &&
React.createElement("div", { className: "overlap", style: { top: getWaveTrueOffset(props.waves, 1), } }, props.children),
props.waves.map((svg, index) => {
const sign = (props.concurrentZDelta ? props.concurrentZDelta : 0);
let zIndex = zIndexBase + index * sign + (svg.reversed ? (sign ? -sign : sign) : sign);
let responsiveHeight = 1 + (1536 - window.innerHeight) / WIDTH_SCALING_FACTOR;
return (React.createElement("div", { key: index, className: "wave", style: { zIndex: zIndex, top: (trueOffsets[index] + (svg.offset * 1 / responsiveHeight)), } },
svg.reversed &&
React.createElement("div", { ref: heightRefs[index], style: {
height: blockHeights[index],
backgroundColor: svg.color,
} }, svg.content &&
React.createElement(svg.content, Object.assign({}, props.wavesProps))),
React.createElement("svg", { ref: svgRefs[index], xmlns: "http://www.w3.org/2000/svg", viewBox: "0 0 1440 320" },
React.createElement("path", { fill: svg.color, fillOpacity: "1", d: svg.path })),
!svg.reversed &&
React.createElement("div", { ref: heightRefs[index], style: {
height: blockHeights[index],
backgroundColor: svg.color,
} }, svg.content &&
React.createElement(svg.content, Object.assign({}, props.wavesProps)))));
})));
}; //# sourceMappingURL=WavesWidget.js.map } // default/src/components/Content/People_Involved/Attributions/Attributions.jsx $fsx.f[131] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const AccordionWidget_1 = $fsx.r(129); const Buttons_1 = $fsx.r(104); class Attributions extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(130);
});
}
render() {
let accordionData = {
backgroundColor: "bg-green-400",
title: "Attributors to Conjugaid",
itemTitleColor: "text-green-800",
items: [
{
title: "Laboratory and Technical Support",
content: `
Contents
- 1 The Team
- 2 The Department of Biochemistry and Molecular Biology, University of Southern Denmark
- 3 Simon Rose, laboratory technician
- 4 Patrick Rosendahl Andreassen, former iGEM-participant
- 5 Thøger Jensen Krogh, former iGEM participant
- 6 Tina Kronborg , academic Assistant at SDU’s Department of Biochemistry and Molecular Biology
- 7 Eric van der Helm and Andreas Porse from SNIPR Biome
- 8 Finn Kirpekar, PhD and associate professor at the Department of Biochemistry and Molecular Biology at the University of Southern Denmark
- 9 The Academy of Talented Youths
- 10 Viktor Hundtofte Mebus, former iGEM participant
- 11 The Team
- 12 Jesper Terp, Odense Katedralskole (a local high-school) and biotechnology-teacher
- 13 Flemming Nielsen , Msc student
- 14 Jakob Møller-Jensen, PhD, associate professor and head of Department of Biochemistry and Molecular Biology
- 15 Martin Worm-Leonhard, forensic chemist and science communicator with UNF
- 16 Jesper Houmann Kattai, biotechnology-teacher at Odense Technical High-school (OTG)
- 17 Students at Aalborg Katedralskole (a high school)
- 18 Biologiformidling
- 19 SciShow, Crash Course and Kurzgesagt
- 20 Visme
- 21 The Team
- 22 Ute Wolff Sönksen , chief physician at Statens Serum Institut (SSI)
- 23 Alexandra Kruse, MD and specialist in pediatric infectious diseases from Doctors Without Borders (MSF)
- 24 Hans Jørn Kolmos, professor, chief physician and Dr.Med.
- 25 Janne Kudsk Klitgaard, PhD, researcher and associate professor at the Department of Clinical Microbiology at the University of Southern Denmark
- 26 Odense University Hospital
- 27 IDA Student Union
- 28 The Team
- 29 Lise Junker Nielsen, outreach Coordinator at the Department of Biochemistry and Molecular Biology
- 30 Jørgen Dinesen (Biotechnology) and Christian Maaløv Andreasen (Physics and chemistry), Svendborg High School and teachers
- 31 UNF Biotech Camps
- 32 The Danish Science Festival
- 33 The Team
- 34 The Municipality of Odense
- 35 The People’s Democratic Festival
- 36 The Team
- 37 Thøger Jensen Krogh , former iGEM SDU participant and creator of the “Best Wiki 2013” (find it in the Judging Handbook...)
- 38 SDU & Bikubenfonden
- 39 IDA Student Union
- 40 Pharmadanmark Student Union
- 41 Ovulaid (iGEM Team Copenhagen) and DTU BioBuilders, fellow Danish iGEM teams
- 42 The Team
- 43 Jakob Møller-Jensen, head of department of Biochemistry and Molecular Biology at the University of Southern Denmark,
- 44 Marianne Holmer, dean of The Faculty of Science at the University of Southern Denmark,
- 45 Henrik Dam, vice-Cancellor at University of Southern Denmark,
- 46 IDA Student Union and Pharmadanmark Student Union
- 47 Our project supervisors
- 48 All former iGEM participants at SDU
- 49 DTU Biobuilders
- 50 Ovulaid, the iGEM Team Copenhagen
- 51 Nina Bjørnskov
- 52 Anders Boe
- 53 Arvid Larsson from the 2019 iGEM team Lund
- 54 Clare Kirkpatrick, assistant Professor at SDU’s Department of Biochemistry and Molecular Biology
- 55 Freja Cecilie Mikkelsen, biomedicine BSc student
- 56 Morten Johansen, Chemistry MSc student
- 57 Maria Victoria Mikkelsen, PhD Fellow at SDU’s Department of Biochemistry and Molecular Biology and former iGEM participant
- 58 Ulrik Stenz Justesen, associate Professor in clinical microbiology at Odense University Hospital
- 59 Megan Fantes, software Engineer Intern at BU SAIL
- 60 Jackie Fantes, MD
- 61 Ellen Gammelmark, biochemistry and molecular biology MSc student and former iGEM participant
- 62 Thomas Fantes, MD
- 63 Sergi Torres Puig, postdoc at SDU’s Department of Biochemistry and Molecular Biology
- 64 Thøger Jensen Krogh former iGEM SDU participant and creator of the “Best Wiki 2013” (find it in the Judging Handbook...)
- 65 Anders Bruhn Arndal Andersen, PhD student at SDU’s Department of Physics, Chemistry and Pharmacy
- 66 Peter Andersen, PE Innovator & VC Investor
- 67 Esther Hvidtfeldt, veterinary medicine student
- 68 Jakob Jersild Nielsen, Computational biomedicine MSc student
- 69 03.26.19 Conjugation & CRISPR
- 70 03.26.19 Conjugation & CRISPR
- 71 03.27.19 Conjugation & CRISPR
- 72 03.30.19 Conjugation & CRISPR
- 73
- 74
- 75 04.01.19 Conjugation & CRISPR
- 76 04.02.19 Target bacteria design / Fluorescens library
- 77 04.03.19 Conjugation & CRISPR
- 78 04.04.19 Conjugation & CRISPR
- 79 04.05.19 Conjugation & CRISPR
- 80 04.14.19 Target bacteria design / Fluorescens library
- 81 04.16.19 Target bacteria design / Fluorescens library
- 82 04.17.19 Target bacteria design / Fluorescens library
- 83 04.19.19 Target bacteria design / Fluorescens library
- 84 04.19.19 Target bacteria design / Fluorescens library
- 85 05.06.19 Conjugation & CRISPR
- 86 05.09.19 Conjugation
- 87 05.10.19 Conjugation
- 88 05.13.19 Conjugation
- 89 05.14.19 Conjugation
- 90 05.15.2019 and 05.16.2019: CRISPR
- 91 05.13.19 Conjugation
- 92 05.14.19 Conjugation
- 93 05.15.2019 and 05.16.2019: CRISPR
- 94 06.09.19 Conjugation
- 95 06.11.19 Conjugation
- 96 06.11.2019 CRISPR
- 97 06.12.19 Conjugation
- 98 06.13.19 Conjugation
- 99 06.15.19 CRISPR
- 100 06.16.19 CRISPR
- 101 06.18.19 Conjugation
- 102 06.18.2019 CRISPR
- 103 06.20.2019 Conjugation
- 104 06.20.2019 CRISPR
- 105 06.21.2019 CRISPR and Target bacteria design / Fluorescens library
- 106 06.22.2019 CRISPR and Target bacteria design / Fluorescens library
- 107 06.23.2019 Conjugation
- 108 06.23.2019 Target bacteria design / Fluorescens library
- 109 06.24.2019 Conjugation
- 110 06.24.2019 Target bacteria design / Fluorescens library
- 111 06.24.2019 CRISPR
- 112 06.25.2019 Target bacteria design / Fluorescens library
- 113 06.26.2019 Target bacteria design / Fluorescens library
- 114 06.27.2019 Target bacteria design / Fluorescens library
- 115 07.01.2019 Target bacteria design / Fluorescens library
- 116 07.02.19 Conjugation
- 117 07.02.2019 CRISPR
- 118 07.03.19 Conjugation
- 119 07.03.2019 CRISPR
- 120 07.04.2019 CRISPR
- 121 07.05.2019 CRISPR
- 122 07.06.2019 CRISPR
- 123 07.07.2019 CRISPR
- 124 07.08.2019 CRISPR
- 125 07.09.2019 CRISPR
- 126 07.10.19 Conjugation
- 127 07.11.2019 Conjugation
- 128 07.11.2019 CRISPR
- 129 07.12.2019 CRISPR
- 130 07.13.2019 CRISPR
- 131 07.13.2019 Conjugation
- 132 07.14.2019 Conjugation
- 133 07.14.2019 CRISPR
- 134 07.15.2019 CRISPR
- 135 07.16.2019 Conjugation
- 136 07.16.2019 CRISPR
- 137 07.17.2019 Conjugation
- 138 07.17.2019 CRISPR
- 139 07.18.2019 Conjugation
- 140 07.18.2019 CRISPR
- 141 07.19.2019 Conjugation
- 142 07.19.2019 CRISPR
- 143 07.20.2019 CRISPR
- 144 07.21.2019 CRISPR
- 145 07.22.19 CRISPR
- 146 07.23.19 CRISPR
- 147 07.24.19 CRISPR
- 148 07.25.19 CRISPR
- 149 07.26.19 CRISPR
- 150 07.28.19 CRISPR
- 151 07.29.19 CRISPR
- 152 07.30.2019 Conjugation
- 153 07.30.19 CRISPR
- 154 07.31.2019 Conjugation
- 155 07.31.19 CRISPR
- 156 08.01.19 CRISPR
- 157 08.02.19 CRISPR
- 158 08.03.19 CRISPR
- 159 08.04.19 CRISPR
- 160 08.05.2019 Conjugation
- 161 08.05.19 CRISPR
- 162 08.06.2019 Conjugation
- 163 08.06.19 CRISPR
- 164 08.07.2019 Conjugation
- 165 08.07.19 CRISPR
- 166 08.08.19 CRISPR
- 167 08.09.19 CRISPR
- 168 08.11.2019 Conjugation
- 169 08.12.2019 Conjugation
- 170 08.12.2019 CRISPR
- 171 08.13.2019 Conjugation
- 172 08.13.2019 CRISPR
- 173 08.14.2019 CRISPR
- 174 08.14.2019 Target bacteria design / Fluorescens library
- 175 08.15.2019 CRISPR
- 176 08.15.2019 Target bacteria design / Fluorescens library
- 177 08.15.2019 Conjugation
- 178 08.16.2019 CRISPR
- 179 08.17.2019 Conjugation
- 180 08.17.2019 CRISPR
- 181 08.17.2019 Conjugation
- 182 08.18.2019 CRISPR
- 183 08.19.2019 Conjugation
- 184 08.19.2019 CRISPR
- 185 08.20.2019 Conjugation
- 186 08.20.2019 CRISPR
- 187 08.20.2019 Target bacteria design / Fluorescens library
- 188 08.21.2019 Target bacteria design / Fluorescens library
- 189 08.22.2019 CRISPR
- 190 08.22.2019 Conjugation
- 191 08.23.2019 CRISPR
- 192 08.23.2019 Conjugation
- 193 08.24.2019 CRISPR
- 194 08.24.2019 Conjugation
- 195 08.25.2019 CRISPR
- 196 08.26.2019 Conjugation
- 197 08.26.2019 CRISPR
- 198 08.28.2019 Conjugation
- 199 08.29.2019 Conjugation
- 200 08.29.2019 CRISPR
- 201 08.30.2019 CRISPR
- 202 08.31.2019 Conjugation
- 203 08.31.2019 CRISPR
- 204 09.01.2019 Conjugation
- 205 09.01.2019 CRISPR
- 206
- 207 09.03.2019 Conjugation
- 208 09.03.2019 CRISPR
- 209 09.04.2019 CRISPR
- 210 09.05.2019 CRISPR
- 211 09.08.2019 CRISPR
- 212 09.09.2019 Conjugation
- 213 09.09.2019 CRISPR
- 214 09.12.2019 CRISPR
- 215 09.13.2019 Conjugation
- 216 09.13.2019 CRISPR
- 217 09.14.2019 CRISPR
- 218 09.15.2019 Conjugation
- 219 09.15.2019 CRISPR
- 220 09.16.2019 CRISPR
- 221 09.17.2019 CRISPR
- 222 09.18.2019 CRISPR
- 223 09.19.2019 Target bacteria design / Fluorescens library
- 224 09.19.2019 CRISPR
- 225 09.20.2019 CRISPR
- 226 09.20.2019 Target bacteria design / Fluorescens library
- 227 09.21.2019 Conjugation
- 228 09.21.2019 CRISPR
- 229 09.17.2019 CRISPR
- 230 09.22.2019 Conjugation
- 231 09.22.2019 CRISPR
- 232 09.23.2019 CRISPR
- 233 09.23.2019 CRISPR
- 234 09.25.2019 CRISPR
- 235 09.26.2019 Conjugation
- 236 09.26.2019 CRISPR
- 237 09.27.2019 CRISPR
- 238 09.28.2019 Conjugation
- 239 09.28.2019 CRISPR
- 240 10.01.19 Conjugation & CRISPR
- 241 10.01.2019 CRISPR
- 242 10.02.2019 CRISPR
- 243 10.03.19 Conjugation & CRISPR
- 244 10.03.2019 CRISPR
- 245 10.04.2019 CRISPR
- 246 10.05.19 Conjugation & CRISPR
- 247 10.05.2019 CRISPR
- 248 10.05.19 Conjugation & CRISPR
- 249 10.06.2019 CRISPR
- 250 10.07.19 Conjugation & CRISPR
- 251 10.07.2019 CRISPR
- 252 10.08.19 Conjugation & CRISPR
- 253 10.08.2019 CRISPR
- 254 10.09.19 Conjugation & CRISPR
- 255 10.09.2019 CRISPR
- 256 10.10.19 Conjugation & CRISPR
- 257 10.11.19 Conjugation & CRISPR
- 258 10.12.19 Conjugation & CRISPR
- 259 10.13.19 Conjugation & CRISPR
- 260 10.13.2019 CRISPR
- 261 10.14.2019 CRISPR
- 262 10.15.2019 CRISPR
- 263 10.15.2019 CRISPR
- 264 10.19.19 Conjugation & CRISPR
- 265 10.20.19 Conjugation & CRISPR
- 266 10.21.19 Conjugation & CRISPR
- 267
The Team
Our team has independently planned and executed our project in the laboratory. Everything from overnight cultures to qPCR’s have been performed by our team members. However, we received support and assistance whenever we were introduced to new and difficult techniques. For that, we would like to thank the following people:
The Department of Biochemistry and Molecular Biology, University of Southern Denmark
For providing us with lab space and equipment.
Simon Rose, laboratory technician
For general lab support, technical assistance and introduction to safety and practicalities in the lab. Simon has especially been of great assistance when introducing us to various difficult techniques in the lab, such as qPCR and purification of chromosomal DNA.
Patrick Rosendahl Andreassen, former iGEM-participant
For troubleshooting and assistance in the lab and in the development of fluorescence microscope.
Thøger Jensen Krogh, former iGEM participant
For troubleshooting when problems arose in the lab.
Tina Kronborg , academic Assistant at SDU’s Department of Biochemistry and Molecular Biology
For assisting us with performing quantitative FACS-analysis and introduction to fluorescent microscopy.
Eric van der Helm and Andreas Porse from SNIPR Biome
For letting us visit them and giving general and technical feedback and advice on our project.
Finn Kirpekar, PhD and associate professor at the Department of Biochemistry and Molecular Biology at the University of Southern Denmark
For advice on conjugation.
The Academy of Talented Youths
For visiting our facility and helping us purify some of our plasmids.
Viktor Hundtofte Mebus, former iGEM participant
For support with quantitative FACS-analysis and data retrieval.
`
},
{
title: "Human Practices",
content: `
Educational Material
The Team
Our team has independently written all text in our e-book, “What’s Up With Antibiotics?”. Furthermore, we have made every illustration in the book ourselves, as well as filmed and edited start-of-chapter introductory videos. By using the online graphical design tool Visme, our team members have designed the entire book. We have also created the teaching guides and laboratory protocols independently. Lastly, we have created evaluation forms and processed these based on the testing of our material on high school students. We would like to thank the following contributors for helping us shape and create our educational material:
Jesper Terp, Odense Katedralskole (a local high-school) and biotechnology-teacher
For collaborating with us, providing us with 800 minutes of educational hours in his class and for input on the content of our educational material.
Flemming Nielsen , Msc student
For creating an educational video about BLAST for “What’s Up With Antibiotics?”.
Jakob Møller-Jensen, PhD, associate professor and head of Department of Biochemistry and Molecular Biology
For appearing in educational videos created for each chapter of “What’s Up With Antibiotics?”.
Martin Worm-Leonhard, forensic chemist and science communicator with UNF
For inspiration and valuable insights to creating educational material for high-school students.
Jesper Houmann Kattai, biotechnology-teacher at Odense Technical High-school (OTG)
For valuable input on the shortcomings regarding microbiology and synthetic biology in the Danish high-school curriculum.
Students at Aalborg Katedralskole (a high school)
For surveying preliminary chapters and reporting opinions on difficulty and readability.
Biologiformidling
For insights into the practical aspects behind the creation of an educational e-book about science.
SciShow, Crash Course and Kurzgesagt
For allowing us to refer to their educational videos throughout the text in our e-book, “What’s Up With Antibiotics?”.
Visme
For discounts on their software, which we used to design our e-book.
Symposium
The Team
Regarding our symposium on antibiotic resistance hosted at the Odense University Hospital, our team has independently planned the entire event. We contacted the four speakers (see below) and arranged every practicality. In collaboration with the hospital, we arranged catering of coffee, tea and snacks and organized live-streaming of the event. We would like to thank the following contributors:
Ute Wolff Sönksen , chief physician at Statens Serum Institut (SSI)
For speaking at our symposium.
Alexandra Kruse, MD and specialist in pediatric infectious diseases from Doctors Without Borders (MSF)
For speaking at our symposium.
Hans Jørn Kolmos, professor, chief physician and Dr.Med.
For speaking at our symposium.
Janne Kudsk Klitgaard, PhD, researcher and associate professor at the Department of Clinical Microbiology at the University of Southern Denmark
For speaking at our symposium.
Odense University Hospital
For providing the facilities, catering and live-streaming services for our symposium on antibiotic resistance.
IDA Student Union
For sponsoring refreshments to the audience.
Public Engagement
The Team
Our team has independently arranged multiple outreach-events and talks throughout our project. We would like to thank the following contributors for providing us with the opportunities to engage in outreach:
Lise Junker Nielsen, outreach Coordinator at the Department of Biochemistry and Molecular Biology
For providing us with opportunities to engage with high school students via multiple outreach-events.
Jørgen Dinesen (Biotechnology) and Christian Maaløv Andreasen (Physics and chemistry), Svendborg High School and teachers
For reaching out and coordinating a day of workshops at our workspaces at SDU.
UNF Biotech Camps
For inviting us to speak at their events.
The Danish Science Festival
For providing us with a booth at their annual event. Thank you all the visitors attending our booth, especially the kids and youngsters who drew their own “Superbacterium”.
Sustainable Development Goals
The Team
Our team has had the UN’s 17 SDGs in mind throughout our entire project, but we would like to thank the following contributors for creating opportunities to engage with the SDGs in our community:
The Municipality of Odense
For hosting an event on the UN’s 17 Sustainable Development Goals and thereby giving us the opportunity to engage within this field.
The People’s Democratic Festival
For hosting their 9th annual People’s Democratic Festival at the island of Bornholm in Denmark. With the festival they created the perfect foundation for debating democracy and opportunities for sustainable development in many ways.
Danish iGEM Weekend
The Team
Our team independently planned and executed the Danish iGEM Weekend in its entirety, but we would like to thank the following contributors for helping us making the meetup a reality:
Thøger Jensen Krogh , former iGEM SDU participant and creator of the “Best Wiki 2013” (find it in the Judging Handbook...)
For hosting a workshop during the Danish iGEM Weekend on creating a successful iGEM-wiki.
SDU & Bikubenfonden
For providing accommodation for all attending teams during the Danish iGEM Weekend.
IDA Student Union
For sponsoring the Danish iGEM Weekend.
Pharmadanmark Student Union
For sponsoring the Danish iGEM Weekend.
Ovulaid (iGEM Team Copenhagen) and DTU BioBuilders, fellow Danish iGEM teams
For participating.
</div `
},
{
title: "Fundraising",
content: `
The Team
Our team has independently written each fundraising application that we’ve sent out. However, we would like to thank the following contributors for assisting and supporting us along the way:
Jakob Møller-Jensen, head of department of Biochemistry and Molecular Biology at the University of Southern Denmark,
For supporting our iGEM project.
Marianne Holmer, dean of The Faculty of Science at the University of Southern Denmark,
For supporting our iGEM project.
Henrik Dam, vice-Cancellor at University of Southern Denmark,
For supporting our iGEM project.
IDA Student Union and Pharmadanmark Student Union
For sponsoring our Danish iGEM Weekend.
Our project supervisors
For proof-reading our fundraising applications and guiding us along the way.
`
},
{
title: "Other General Support",
content: `
All former iGEM participants at SDU
For attending our preliminary presentation and providing us with valuable feedback before the Giant Jamboree.
DTU Biobuilders
For hosting their 7th annual BioBrick Tutorial and for participating in our Danish iGEM Weekend.
Ovulaid, the iGEM Team Copenhagen
For hosting this year’s Nordic iGEM Conference and for participating in our Danish iGEM Weekend.
Nina Bjørnskov
For giving us the opportunity to promote our project by allowing us to take over the Faculty of Science’s Instagram page (@sduscience) for a week.
Anders Boe
For creating our final team video.
Arvid Larsson from the 2019 iGEM team Lund
For providing valuable advice and support in designing our modeling system.
Clare Kirkpatrick, assistant Professor at SDU’s Department of Biochemistry and Molecular Biology
For providing feedback on our wiki-text.
Freja Cecilie Mikkelsen, biomedicine BSc student
For providing feedback on our wiki-text.
Morten Johansen, Chemistry MSc student
For providing feedback on our wiki-text.
Maria Victoria Mikkelsen, PhD Fellow at SDU’s Department of Biochemistry and Molecular Biology and former iGEM participant
For providing feedback on our wiki-text.
Ulrik Stenz Justesen, associate Professor in clinical microbiology at Odense University Hospital
For providing feedback on our wiki-text.
Megan Fantes, software Engineer Intern at BU SAIL
For providing feedback on our wiki-text (and for her ongoing support and cheering-on of our team).
Jackie Fantes, MD
For providing feedback on our wiki-text.
Ellen Gammelmark, biochemistry and molecular biology MSc student and former iGEM participant
For providing feedback on our wiki-text.
Thomas Fantes, MD
For providing feedback on our wiki-text.
Sergi Torres Puig, postdoc at SDU’s Department of Biochemistry and Molecular Biology
For providing feedback on our wiki-text.
Thøger Jensen Krogh former iGEM SDU participant and creator of the “Best Wiki 2013” (find it in the Judging Handbook...)
For providing feedback on our wiki-text and layout.
Anders Bruhn Arndal Andersen, PhD student at SDU’s Department of Physics, Chemistry and Pharmacy
For providing feedback on our wiki-text.
Peter Andersen, PE Innovator & VC Investor
For providing feedback on our wiki-text.
Esther Hvidtfeldt, veterinary medicine student
For providing feedback on our wiki-text.
Jakob Jersild Nielsen, Computational biomedicine MSc student
For providing valuable guidance in constructing our conjugation models
`
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "attributions" },
React.createElement("div", { className: "content pb-20" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Attributions"),
React.createElement("div", { className: "under-rubrik" }, "Thank you\u2019s"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "Our team began our project by brainstorming project ideas in January 2019. We had many initial ideas, but ultimately chose to work on Conjugaid - an idea thought up solely by our team-members. "),
React.createElement("p", null, "In March 2019 we begun working in the lab after an introductory safety and practicalities course. We have accomplished many things on our own, including the formulation, design and execution of our laboratory experiments as well as the writing, designing and planning of our educational material and lesson plan (e-book, illustrations, teaching guides, laboratory protocols and evaluations included). Additionally, we have independently managed and dealt with all communications regarding Integrated Human Practices, Fundraising and promotion of our project. All work within modelling and data processing has been carried out by Conjugaid team members."),
React.createElement("p", null, "However, there are limitations to what one can accomplish alone, which is why our team would like to say thank you to everyone involved in our project. Keep reading to get to know the people who helped us make Conjugaid a reality."))),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionData }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Attributions = Attributions; //# sourceMappingURL=Attributions.js.map } // default/src/components/widgets/AccordionWidget.jsx $fsx.f[129] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const _ = $fsx.r(44); const react_scroll_1 = $fsx.r(128); class AccordionWidget extends React.Component {
constructor(props) {
super(props);
}
scrollToTop(e) {
let target = e.currentTarget;
let top = target.nextElementSibling.getBoundingClientRect().top;
let menu = document.querySelector(".top-menu").getBoundingClientRect();
let delta = top - menu.height;
if (delta > 0) {
}
react_scroll_1.animateScroll.scrollTo(window.scrollY + delta - 50);
}
render() {
let id = _.uniqueId();
return (React.createElement("nav", { className: "accordion arrows" },
React.createElement("header", { className: "box ".concat(this.props.model.backgroundColor) },
React.createElement("label", { htmlFor: `acc-close${id}`, className: "box-title" }, this.props.model.title)),
this.props.model.items.map((element, index) => {
return (React.createElement("div", { key: index },
React.createElement("input", { type: "radio", name: `accordion${id}`, id: `${id}${index}`, onClick: this.scrollToTop }),
React.createElement("section", { className: "box" },
React.createElement("label", { className: "box-title ".concat(this.props.model.itemTitleColor), htmlFor: `${id}${index}` }, element.title),
React.createElement("label", { className: "box-close", htmlFor: `acc-close${id}` }),
React.createElement("div", { className: "box-content", dangerouslySetInnerHTML: { __html: element.content } }))));
}),
React.createElement("input", { type: "radio", name: `accordion${id}`, id: `acc-close${id}` })));
}
} exports.AccordionWidget = AccordionWidget;
//# sourceMappingURL=AccordionWidget.js.map } // default/src/components/Content/Human_Practices/Public_Engagement/PublicEngagement.jsx $fsx.f[171] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const AccordionWidget_1 = $fsx.r(129); const WavesWidget_1 = $fsx.r(110); const Buttons_1 = $fsx.r(104); const rodal_1 = $fsx.r(132); const nuka_carousel_1 = $fsx.r(168); const react_animate_on_scroll_1 = $fsx.r(169); const topWaves = [
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,96L48,128C96,160,192,224,288,240C384,256,480,224,576,213.3C672,203,768,213,864,229.3C960,245,1056,267,1152,277.3C1248,288,1344,288,1392,288L1440,288L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
}
]; class PublicEngagement extends React.Component {
constructor(props) {
super(props);
this.state = { visible: false, visibleTwo: false, visibleThree: false, carouselHeight: 0, carouselTwoHeight: 0 };
this.carousel = React.createRef();
this.carouselTwo = React.createRef();
window.document.title = `${props.title} | SDU-Denmark`;
}
show(obj) {
document.body.classList.add("rodal-open");
this.setState(obj);
}
hide(obj) {
document.body.classList.remove("rodal-open");
this.setState(obj);
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(170);
});
}
componentDidMount() {
let carousel = this.carousel.current;
if (carousel) {
let constant = 0.5625;
let parent = carousel.frame.parentElement;
let parentRect = parent.getBoundingClientRect();
let newheight = parentRect.width * constant;
this.setState({
carouselHeight: newheight
});
}
let carouselTwo = this.carouselTwo.current;
if (carouselTwo) {
let constant = 0.5625;
let parent = carouselTwo.frame.parentElement;
let parentRect = parent.getBoundingClientRect();
let newheight = parentRect.width * constant;
this.setState({
carouselTwoHeight: newheight
});
}
}
render() {
let accordionData = {
backgroundColor: "bg-pink-600",
title: "Click on an event to learn more",
itemTitleColor: "text-yellow-600",
items: [
{
title: "SDU Biotech Camp",
content: `
A few weeks after the beginning of our project, we had the opportunity to talk about iGEM and our project to a group of high school students visiting University of Southern Denmark for a weekend during the SDU Biotech Camp. Here, we were asked many curious questions, especially about the prospects of our project being applied clinically. Our discussions with them helped us specify our own project design.
<img id="box-img" src="
"></img>
},
{
title: 'Svendborg High School',
content: `
Students from <a href="https://www.svendborg-gym.dk" target="_blank">Svendborg Gymnasium</a> came to visit us at SDU. We talked about iGEM and guided the students through a tour of our laboratory. Finally, we hosted a workshop on the iGEM BioBrick system.
For the workshop, we had come up with five different case-scenarios that needed to be solved by creating biobricks from a range of parts in the registry. The scenarios were inspired by different SDG’s and included: plastic pollution, CO2 in the atmosphere, pharmaceuticals in wastewater, iron deficiency in humans and lactose intolerance. The students worked in teams to choose the appropriate parts needed for their solution onto a standard plasmid template provided by us.
<img id="box-img" src="
"></img>
In the evaluation, we could see that 18 students were more likely to pursue a university degree in science, and all students got a better understanding of synthetic biology.
<img id="box-img" src="
"></img>
<img id="box-img" src="
"></img>
`
},
{
title: "UNF Biotech Camp",
content: `
We were invited to give a joined talk along with<a href="https://2019.igem.org/Team:UCopenhagen" target="_blank"> Ovulaid (iGEM Team Copenhagen 2019)</a> and <a href="https://2019.igem.org/Team:DTU-Denmark" target="_blank">DTU Biobuilders (iGEM Team DTU 2019)</a> at the annual Biotech Science Camp hosted by The Danish Youth Association of Science (UNF). Here, each team presented their projects to high school students.
<img src="
"> </img>
`
},
{
title: "The Academy of Talented Youths",
content: `
We were invited to host an evening of talks and laboratory-work for high school students from <a>The Academy of Talented Youths</a>. Here, we gave a talk about iGEM and synthetic biology. Furthermore, we helped the students in the lab as they purified our plasmids, which we have later used in our laboratory work. This indeed helped our team, as the end of the competition was drawing near.
<img src="
"> </img>
`
},
{
title: "Nykøbing F. High School",
content: `
We gave a talk about iGEM and synthetic biology to high school students from Nykøbing Falster. We also discussed the ethical considerations around working with synthetic biology.
`
}
]
};
let accordionData2 = {
backgroundColor: "bg-pink-600",
title: "References",
itemTitleColor: "text-yellow-600",
items: [
{
title: "Education & Public Engagement References",
content: `
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/25768880" target="_blank" class="references">Obrist, R. (2015). Only conflicts of interest? Swiss Med Wkly, 145:w14120. </a>
- <a href="https://www.who.int/antimicrobial-resistance/publications/global-action-plan/en" target="_blank" class="references">World Health Organization (2015). Global action plan on antimicrobial resistance. </a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "PE" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Education & Public Engagement"),
React.createElement("div", { className: "under-rubrik" }, "SDG no. 4: Quality Education"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" },
" In a time where perception of scientific credibility is changing in the eyes of the general public ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/25768880", target: "_blank" }, "Obrist, R. (2015). Only conflicts of interest? Swiss Med Wkly, 145:w14120. "))),
", it is more important than ever to maintain the integrity of the scientific community. Without the support of governments and the public, we as scientists cannot combat problems like antibiotic resistance. One way to help maintain this integrity is to engage the general public by opening the scientific community. We have worked hard within education and public engagement in order to do this successfully - keep reading to learn more. ")))),
React.createElement(react_animate_on_scroll_1.default, { animateIn: "fadeIn" },
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row", id: "no-bottom" },
React.createElement("h2", { className: "darker-border" }, "Education"),
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "darker-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CEducation is the most powerful weapon which you can use to change the world.\u201D "),
React.createElement("p", { className: "quotemaster" }, "- Nelson Mandela, South African anti-apartheid revolutionary"),
React.createElement("div", { className: "darker-line" }, " ")),
React.createElement("h3", null, "Educational Material for High School Students"),
React.createElement("p", null,
React.createElement("span", { className: "italic" }, " \u201CWell, try to tell me what has happened since you arrived back home from Thailand\u201D"),
" said the Doctor. This is the beginning of the story of \u201CWhat\u2019s Up With Antibiotics?\u201D, the open-source, case-oriented e-book produced by our team. The e-book is a part of a lesson-plan created by our team in collaboration with Danish high school students and teachers."),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "
" })),
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "
" })),
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "
" }))),
React.createElement("p", null,
"The e-book is supported by teaching manuals and laboratory protocols developed by our team. It is aimed at Danish high school students majoring in biotechnology, and it supports the present curriculum. After surveying students, we learned that electronic materials such as videos and illustrations is beneficial for their learning experience, which guided us in our decision-making process. You can view the e-book in Danish ",
React.createElement("a", { target: "_blank", href: "https://static.igem.org/mediawiki/2019/d/d1/T--SDU-Denmark--ebog_dansk.pdf" }, "here"),
" or read an English excerpt of the first two chapters ",
React.createElement("a", { target: "_blank", href: "https://static.igem.org/mediawiki/2019/7/74/T--SDU-Denmark--ebog_engelsk.pdf" }, "here"),
"."),
React.createElement("img", { id: "", className: "rounded-lg my-10", src: "
" }),
React.createElement("p", null,
"According to the first objective of the WHO Global Action Plan on Antimicrobial Resistance ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.who.int/antimicrobial-resistance/publications/global-action-plan/en/", target: "_blank" }, "World Health Organization (2015). Global action plan on antimicrobial resistance. "))),
", awareness about antibiotic resistance through public communication and education is needed. This requires continuous effort, and it was therefore important to us that our educational material is made sustainable and thus accessible for future generations. Therefore, \u201CWhat\u2019s Up With Antibiotics?\u201D is open-source, electronic and easily adaptable by educators since it is accompanied by teaching guides and easy-to-use laboratory protocols."),
React.createElement("p", null, "We decided to frame the e-book as a clinical case at a hospital\u2019s Department of Microbiology, in which the students act as molecular biologists diagnosing and treating Peter, a patient suffering from a resistant bacterial infection. "),
React.createElement("p", null, "The e-book frequently refers to educational videos and covers the following topics:"),
React.createElement("ul", null,
React.createElement("li", null, "Bacteriology"),
React.createElement("li", null, "DNA sequencing"),
React.createElement("li", null, "DNA Blasting"),
React.createElement("li", null, "Antibiotics and antibiotic resistance"),
React.createElement("li", null, "The CRISPR-technology and the bioethics related to it"),
React.createElement("li", null, "Creation of scientific posters"),
React.createElement("li", null, "Constructive feedback-methods")),
React.createElement("p", null, "Using the accompanying laboratory protocols, the students perform PCR and gel electrophoresis as well as two digital exercises elaborating on BLAST and CRISPR. The lesson-plan is rounded off with the students creating a scientific poster of their discoveries, presenting it at a poster session and providing feedback to their peers. "),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: " ", src: "
" })),
React.createElement("p", null,
"When creating our educational material, we have been inspired by the ",
React.createElement("a", { href: "https://lesley.edu/article/empowering-students-the-5e-model-explained", target: "_blank" }, "5E Teaching Model"),
", which allows students to learn new concepts through a cycle of five steps. You can read more about the five steps and our use of them here."),
React.createElement("ol", null,
React.createElement("li", null,
React.createElement("p", null,
React.createElement("span", null, "Engage"),
": The concept of interest is introduced to the students. Equal academic level amongst the students was ensured by giving a relatable introduction to microbiology (through the story of Peter) in the first parts of \u201CWhat\u2019s Up With Antibiotics?\u201D")),
React.createElement("li", null,
React.createElement("p", null,
React.createElement("span", null, "Explore"),
": The students extend their knowledge through hands-on learning. We introduced students to the scientific methods behind microbiology through laboratory exercises.")),
React.createElement("li", null,
React.createElement("p", null,
React.createElement("span", null, "Explain"),
": The teacher aids the students in understanding their work during the Explore-phase. The laboratory-results was explained to and discussed with the students (eg. the significance of the bands on their gels).")),
React.createElement("li", null,
React.createElement("p", null,
React.createElement("span", null, "Elaborate"),
": In order for the students to gain a deeper understanding of their work, they are asked to elaborate their obtained knowledge by creating a scientific poster about the course and presenting it to their peers.")),
React.createElement("li", null,
React.createElement("p", null,
React.createElement("span", null, "Evaluate"),
": The students perform self- and peer-assessment of their scientific presentations using the BIO-feedback model, which is presented to them in the last chapter of \u201CWhat\u2019s Up With Antibiotics?\u201D"))),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null, "The entire lesson plan was fully tested on students from the Danish high school Odense Katedralskole. These students are majoring in biotechnology and was our primary test group. We tested our materials over a period of four weeks. "),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null,
"Click here to read some of the feedback we received from the ",
React.createElement("button", { className: "text-orange-600", onClick: this.show.bind(this, { visible: true }) }, "students"),
"."),
React.createElement(rodal_1.default, { closeOnEsc: true, customStyles: {
width: "80vw",
height: "80vh"
}, visible: this.state.visible, onClose: this.hide.bind(this, { visible: false }) },
React.createElement("div", { className: "rodal-box" },
React.createElement("div", { className: "border-solid border-gray-700 border-b " },
React.createElement("h3", null, "Student feedback on our teaching resources")),
React.createElement("div", { className: "italic" },
React.createElement("p", null,
"\u201CIt\u2019s nice to watch ",
React.createElement("span", { className: "font-bold" }, "videos"),
" once in a while, since I can get tired of reading.\u201D"),
React.createElement("p", null,
"\u201CI think the level of difficulty was good. Some things were complicated but ",
React.createElement("span", { className: "font-bold" }, "we need challenges"),
". I think it was a really good book and a really good course. It has been ",
React.createElement("span", null, "educative "),
" and most of the class liked it.\u201D"),
React.createElement("p", null,
"\u201CNow that I\u2019ve gained knowledge of something new, ",
React.createElement("span", { className: "font-bold" }, "I can reflect upon it"),
". Maybe I have an idea that\u2019s worth working on\u2026\u201D"),
React.createElement("p", null,
"\u201CI really enjoyed the introductions to each chapters, ",
React.createElement("span", { className: "font-bold" }, "the ones about Peter"),
". They made it easy to grasp the situation and made me understand the material better.\u201D"),
React.createElement("p", null,
"\u201C[...] I also really liked that you made ",
React.createElement("span", { className: "font-bold" }, "glossaries"),
", so I could actually understand what I was reading about without having to google everything all the time.\u201D")))),
React.createElement("p", null, "From creating educational materials for high school students, we have gathered our experiences into a 10-step guide:"),
React.createElement("h3", null, "10 steps to creating great educational material"),
React.createElement("ol", null,
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Find out your target group"),
React.createElement("p", { className: "italic" }, "It is imperative when creating educational material to know who your students are. Teaching children in elementary school about microbes requires something completely different than teaching high schoolers does.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Talk to your target group (and their teachers)"),
React.createElement("p", { className: "italic" }, "Before you can develop great materials, you need to know the needs of the recipients. What do they lack, and what do they already have in abundance? Your way forward is to fill out the gaps.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Tell a story"),
React.createElement("p", { className: "italic" }, "One important rule of thumb about education is that learning is eased through storytelling. We taught antibiotic resistance through the story of Peter - how will you tell yours?")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Plan out everything in detail"),
React.createElement("p", { className: "italic" }, "Before you can start writing and designing your educational material, you need a solid plan. How many subjects do you wish to cover - and what should they be? What kind of illustrations do you need? Sort everything into chapters and write down what they should contain. You could be inspired by different teaching and learning models, like we did with the 5E Teaching Model.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Write, write and rewrite..."),
React.createElement("p", { className: "italic" }, "Start writing your materials! And when you think you are done, look at it again - is it really done? Great! Now get someone else to read and comment on it, and then rewrite the whole thing to perfection.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Make it look stunning!"),
React.createElement("p", { className: "italic" }, "This is where you start taking your educational material to the next level. Does someone on your team love drawing? Ask them to create the illustrations and get designing - the options are endless, so have fun with it!")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Finishing touches"),
React.createElement("p", { className: "italic" }, "When your educational material is all done - written, illustrated and designed - forward it to a group of people and get their final comments on it. Make sure to implement these comments before testing your material.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, " Survey before..."),
React.createElement("p", { className: "italic" }, "Congratulations! You have now created your very own educational material - that is amazing. All that is left now is to test it out on your target group and find out how they like it. But before you can test your material and evaluate it, you need to survey your test group. That way, you can compare how your educational material has directly affected the people using it.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Put it to the test"),
React.createElement("p", { className: "italic" }, "Go on - do it! You have nothing to worry about, you have worked so hard and put so much effort into it, we are sure your educational material will do amazingly with your test group.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "\u2026 And evaluate after!"),
React.createElement("p", { className: "italic" }, "This is the last, but definitely not least important, step of this recipe. You have created this amazing educational resource, and you want more people to notice and use it, right? In order for them to do so, they need to see results. Ask your test group any questions that might be relevant for you to properly evaluate how your educational material did in practice."))),
React.createElement("p", null,
"With \u201CWhat\u2019s Up With Antibiotics?\u201D, we aim to inform young minds about antibiotic resistance and synthetic biology. Teaching high school students about the opportunities that lie within these fields, we potentially help shaping the scientists of tomorrow. However, ",
React.createElement("a", { href: "https://sustainabledevelopment.un.org/sdg4", target: "_blank" }, "SDG no. 4"),
" about quality education focuses on ensuring ",
React.createElement("span", { className: "italic" },
"\u201Cinclusive and equitable quality education and promote lifelong learning opportunities",
React.createElement("span", { className: "font-bold" }, " for all"),
"\u201D"),
" and so do we. Thus, our project focuses not only on educating students, but also legislators as well as children and adults of the general public."),
React.createElement("h3", null, "Fluorescence Microscopy for All"),
React.createElement("p", null,
" ",
React.createElement("img", { id: "", className: "icon", src: "
" }),
"For high schools using our educational material in the future, we have assisted in the creation of an open-source recipe for a 3D-printable microscope-attachment. This unique attachment is easy to use by anyone, and provides light microscopes with the properties of fluorescence microscopes. The attachment can be printed by anyone with access to a 3D-printer, and is therefore a great addition to high schools or universities who do not have access to state-of-the-art fluorescence microscopes. Furthermore, it can be incorporated in school systems of low- and middle-income countries, bringing synthetic biology across societal borders and ensuring quality educational opportunities for all. You can learn more about how this system works and find the recipe needed to 3D-print your own free of charge ",
React.createElement("button", { className: "text-orange-600", onClick: this.show.bind(this, { visibleTwo: true }) }, " here"),
"."),
React.createElement(rodal_1.default, { closeOnEsc: true, customStyles: {
width: "80vw",
height: "80vh"
}, visible: this.state.visibleTwo, onClose: this.hide.bind(this, { visibleTwo: false }) },
React.createElement("div", { className: "rodal-box" },
React.createElement("div", { className: "border-solid border-gray-700 border-b " },
React.createElement("h3", null, "Fluorescence Microscopy for All")),
React.createElement("p", null, "The 3D printed microscope attachment uses the basic principles of fluorescence microscopy. Here, the light from an attachable flashlight, which can produce white light, will go through an excitation filter, that only allows the excitation wavelength to pass. Then, the light will be reflected onto the sample, where the emission will then pass through the dichroic mirror and emission filter, and a picture can be taken with a laboratory digital camera. The attachment contains a house, a phone station and a compartment with three cubes that can be customized to the different fluorescence proteins. The cubes can be pushed through the house for easy and quick shift between filters. Each compartment contains a chute for an excitation filter, a dichroic mirror and an emission filter. These filters can be chosen specifically for the type of fluorescence protein in question. Conjugaid has used this microscope for identifying GFP in our samples using Roscolux filters. For GFP, we chose the filters #85 for excitation and #389 for emission based on the known fluorescence spectrum for GFP."),
React.createElement("p", null,
"You can download and use the 3D-print recipe ",
React.createElement("a", { href: "https://www.thingiverse.com/thing:3886674?fbclid=IwAR1Ar6tExUvr_tcrldw_ySLO9IMj5JY3oANDRgg5PRNqHCSaJF9qWjdUpL4", target: "_blank" }, "here"),
"."))),
React.createElement("h3", null, "What's Next?"),
React.createElement("p", null, "With Conjugaid, our team dared to dream big. We dreamed of resolving antibiotic resistance both in the laboratory and by making knowledge available throughout society. After finishing our iGEM project, these dreams have only grown bigger."),
React.createElement("p", null,
"Following our work with\u201CWhat\u2019s Up With Antibiotics?\u201D, we have received an offer to develop it further after iGEM 2019. When speaking at our symposium, Ute Wolff S\u00F6nksen let us know of ",
React.createElement("a", { href: "https://e-bug.eu/", target: "_blank" }, " E-bug "),
". E-bug is an online educational hub about hygiene, microbiology and antibiotics for children and young people and is distributed throughout Europe in 22 different languages. It is partly funded by The Directorate-General for Health and Consumers (DG SANCO) of the European Commission. E-bug offered to translate \u201CWhat\u2019s Up With Antibiotics?\u201D and distribute it on their national and international sites. This is an amazing opportunity to develop on one of Conjugaid\u2019s biggest goals; increasing public awareness about antibiotic resistance. After attending the Giant Jamboree, we will prepare \u201CWhat\u2019s up with antibiotics?\u201D for international distribution by revising it throughout."),
React.createElement("p", null, "Furthermore, we have been contacted by high school teachers requesting similar courses to the one we arranged with Odense Katedralskole. Thus, we will be working closely with a technical high school in Fredericia when iGEM 2019 is over, in order to provide them with a fun and informative few weeks of antibiotic-education. When doing this we will be implementing the 3D-printable microscope-attachment mentioned earlier. "),
React.createElement("p", { id: "no-bottom" }, "After Conjugaid\u2019s birth and development into maturity, our dream is that our project can help shape the scientists and world heroes of tomorrow - because who knows; maybe the next Alexander Fleming will be inspired by \u201CWhat\u2019s Up With Antibiotics?\u201D in the near future.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 }))),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("h2", { className: "whiter-border" }, "Public Engagement"),
React.createElement("h3", null, "The People\u2019s Democratic Festival"),
React.createElement("p", null,
"After attending the municipal meeting in April and having multiple discussions concerning local implementation of the SDGs, it remained clear for us that there is uncertainty regarding the use of synthetic biology amongst the general public. Therefore, we decided to put effort into changing that fact by participating in Denmark\u2019s annual festival about celebrating democracy - ",
React.createElement("a", { href: "https://folkemoedet.dk/en/", target: "_blank" }, "The People\u2019s Democratic Festival"),
". "),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null, "The festival facilitates people and politicians coming together to debate current political issues. It is the aim of the festival to create bridges between the general public and politicians and researchers - the perfect setting to put synthetic biology on the political agenda. We set out to initiate debates concerning how synthetic biology can be applied to fulfil the SDGs as well as the ethical considerations in that process."),
React.createElement("p", null, "At the festival, we had a range of discussions and talks about the use of synthetic biology and the increasing threat of antibiotic resistance with people from different parts of society - from the general public to politicians, researchers and communications experts. Amongst others was the current Prime Minister of Denmark, Mette Frederiksen, DNA-researcher Eske Willerslev as well as EU-Commissioner Margrethe Vestager. "),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null,
"Noteworthy is our talk with Christine Antorini, former Danish Minister for Education and current Director of ",
React.createElement("a", { href: "https://life.dk/om-life/english/", target: "_blank" }, "LIFE "),
"- the Novo Nordisk Foundation\u2019s new learning centre for children and adolescents. Antorini stated the necessity of educational material addressing innovative solutions to world problems (eg. through synthetic biology). In continuation hereof, she further stated the cruciality of the format being case-based to attract the students\u2019 interest. "),
React.createElement("p", null, "Are you interested in hearing more from The People\u2019s Democratic Festival? Then hit \u2018play\u2019 on the video below."),
React.createElement("video", { controls: true },
React.createElement("source", { src: "https://static.igem.org/mediawiki/2019/6/68/T--SDU-Denmark--fm19-education.mp4", type: "video/mp4" })),
React.createElement("p", null, "After talking with leading politicians and researchers, it stood clear to us that the use of synthetic biology is a rather unknown topic for the majority of the public. This amplifies the importance of spreading knowledge about synthetic biology to all groups of society, which definitely strengthened our future work within outreach and public engagement. "),
React.createElement("p", null, "Besides engaging Danish adults and legislators in the debate about antibiotic resistance, we have worked hard at doing the same with the Danish youth. Keep reading to learn more about our public engagement efforts with high school students."),
React.createElement("h3", null, "Public Engagement of High School Students"),
React.createElement("p", null, "To build a foundation from which young minds can create innovative solutions to problems such as antibiotic resistance, we have been in close contact with students from all over Denmark. Read more about the individual outreach events in the foldouts below:"),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionData })),
React.createElement("h3", null, "The Danish Science Festival"),
React.createElement("p", null,
React.createElement("a", { target: "_blank", href: "http://forsk.dk/the-danish-science-festival" }, "The Danish Science Festival"),
" is organized by The Ministry of Higher Education and Science in Denmark, and it aims at enhancing the relationship between scientists and the general public by supporting public scientific engagement. This was the perfect surroundings to engage the people in synthetic biology and the ethical considerations behind it."),
React.createElement("p", null, "We participated by showcasing our project idea. Hereby, we addressed the importance of solving global issues by combining research and innovation. We had two primary activities for the attendees at our workshop; one for the children and one for adolescents and adults. The activity aimed at children was called \u201CMy super-bacteria\u201D. Here, children designed their own genetically engineered super-bacterias with different functionalities - we loved to see their curiosity!"),
React.createElement("p", null,
"You can see a selection of the drawings ",
React.createElement("button", { className: "text-orange-600", onClick: this.show.bind(this, { visibleThree: true }) }, "here"),
"."),
React.createElement(rodal_1.default, { closeOnEsc: true, customStyles: {
width: "100vw",
height: "80vh"
}, visible: this.state.visibleThree, onClose: this.hide.bind(this, { visibleThree: false }) },
React.createElement("div", { className: "rodal-box" },
React.createElement("div", { className: "border-solid border-gray-700 border-b " },
React.createElement("h3", null, "Super Bacteria Drawings done by Children at the Danish Science Festival")),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("p", { className: "italic" }, "Bj\u00F8rk wanted his super-bacterium to eat plastic."),
React.createElement("img", { src: "
" })),
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("p", { className: "italic" }, "Clara, 9 years old, wanted her super-bacterium to make her sushi for dinner."),
React.createElement("img", { src: "
" })),
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("p", { className: "italic" }, "Thea, 13 years old, made her super-bacterium into an LGBTQ+ cow radiating rainbow-love."),
React.createElement("img", { src: "
" })),
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("p", { className: "italic" }, "Kim, 11 years old, wants his super-bacterium to eat micro-plastic and turn it into Christmas decorations"),
React.createElement("img", { src: "
" }))))),
React.createElement("p", null, "For adolescents and adults, we had prepared a more challenging activity explaining the mechanism of CRISPR/Cas. The participants matched a sgRNA in a Cas-protein to a sequence found in DNA-strands. Hereby, we addressed how the CRISPR/Cas-system is applied in our project design. While parents talked to us about genetic engineering and our project, their children designed imaginative super-bacterias. Thus, we managed to engage in a multi-way discussion about synthetic biology with people of all ages. It was a wonderful way to have our project design reviewed early in the process."),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("h3", null, "Symposium"),
React.createElement("p", null, "Our interest in antibiotic resistance lead us to arrange a symposium on the topic for our peers both inside and outside of iGEM. The symposium took place at our local hospital, Odense University Hospital. We gave a project presentation ourselves, and we were followed by interesting talks by four speakers: "),
React.createElement("ul", null,
React.createElement("li", null,
"Chief Physician at Statens Serum Institut (SSI), ",
React.createElement("span", { className: "font-bold" }, "Ute Wolff S\u00F6nksen"),
", gave a talk about the status quo of antibiotic resistance in Denmark."),
React.createElement("li", null,
"MD and specialist in pediatric infectious diseases, ",
React.createElement("span", { className: "font-bold" }, "Alexandra Kruse"),
" from Doctors Without Borders (MSF), gave a talk on how antibiotic resistance affects their work in the field."),
React.createElement("li", null,
"Professor and MD from the Center for Translational Microbiology, ",
React.createElement("span", { className: "font-bold" }, "Hans J\u00F8rn Kolmos"),
", gave a history lesson on MRSA development in Denmark."),
React.createElement("li", null,
"PhD and Associate Professor at the Department of Clinical Microbiology and the Center for Translational Microbiology at the University of Southern Denmark, ",
React.createElement("span", { className: "font-bold" }, "Janne Kudsk Klitgaard"),
", gave a presentation on her research with cannabinoids in combinatorial treatment of resistant bacteria.")),
React.createElement("p", null,
"We live-streamed the symposium and made it publicly available on YouTube afterwards. ",
React.createElement("a", { target: "_blank", href: "https://youtu.be/-ji_ZAPKcHE" }, "Click here to watch it yourself"),
"."),
React.createElement(nuka_carousel_1.default, { ref: this.carouselTwo, className: "mx-auto my-10", width: "80%", dragging: true, style: {
height: this.state.carouselTwoHeight,
} },
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" })),
React.createElement("h3", null, "Engagement in Social Media"),
React.createElement("p", null, "The use of social media is widespread globally as well as across generations. Thus, we were engaging actively with people on social media. "),
React.createElement("p", null,
"Having more than 500 followers on a social media platform is like having a full auditorium of individuals listening to you. Therefore, ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4520913", target: "_blank" }, "Tan, S. Y. and Tatsumura, Y. (2015). Alexander Fleming (1881-1950): Discoverer of penicillin. Singapore Med J. 2015; 56(7): 366-367."))),
"social media has the potential to be an asset for scientists to reach out to the public and spread their messages. We have tried to do so in regard to Conjugaid, by continuously reaching out to our 1,700+ followers spread out on three social media platforms. We even had a Danish Member of Parliament, Sofie Carsten Nielsen, post a photo of us on her story to her 4,000+ followers. "),
React.createElement("p", null,
"Our activities on social media have also helped bring along collaborations between ourselves and other iGEM teams. Through group-chats with other iGEM teams on Instagram, we have discussed our individual projects, started collaborations and helped troubleshoot issues for different teams. For instance, our ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Collaborations", target: "_blank" }, "collaboration"),
" with ",
React.createElement("a", { href: "https://2019.igem.org/Team:Amazonas-Brazil", target: "_blank" }, "Amazonas BeliE.V.E"),
" has been established throughout conversations on Instagram."),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null,
"Finally, we have been active in blogging. ",
React.createElement("a", { href: "https://2015.igem.org/Team:Yale/practices", target: "blank" }, "The Team Yale LGBTQ+ survey in 2015"),
" showed that more than half of the respondents wanted iGEM to improve the discussion on how identity impacts one\u2019s scientific career. Inspired by this, the ",
React.createElement("a", { href: "https://www.chemistryworld.com/opinion/no-sexuality-please-were-scientists/7197.article", target: "_blank" }, "Chemistry World opinion \u201CNo sexuality please, we\u2019re scientists\u201D"),
", and the ",
React.createElement("a", { target: "_blank", href: "https://www.nytimes.com/2015/09/05/opinion/manil-suri-why-is-science-so-straight.html" }, "New York Times opinion \u201CWhy is science so straight?\u201D"),
", a team-member wrote a ",
React.createElement("a", { href: "https://blog.igem.org/blog/2019/10/16/a-letter-to-my-role-model?fbclid=IwAR05hQ-U9JQPxB6zaITaNqVMu9anZiullaCq8eXxgj49JGyTh25cyXag8Ko", target: "_blank" }, "two-piece post"),
" for the iGEM blog with Landon Getz from the Diversity and Inclusion Committee on why LGBTQ+ visibility in STEM fields is important. Improving the iGEM Competition is an iterative process requiring teams, PI\u2019s, and iGEM officials to look inwards. By posting on the iGEM blog we hope to reach the people who need it and their peers, thus assisting this process."),
React.createElement("p", null, "Click \u2018Next\u2019 to learn how future iGEM teams can take our project even further."))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.PublicEngagement = PublicEngagement; //# sourceMappingURL=PublicEngagement.js.map } // default/src/components/Content/People_Involved/Partners/partners.jsx $fsx.f[173] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); class PartnersSponsors extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(172);
});
}
render() {
return (React.createElement("div", { id: "sponsors" },
React.createElement("div", { className: "content pb-20" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Sponsors & Partners"),
React.createElement("div", { className: "under-rubrik" }, "Without you, no us"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "We would like to say a special thank you to all of our sponsors - without them, Conjugaid would never have become what it is today."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.sdu.dk/en/", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "The Faculty of Science at University of Southern Denmark"),
React.createElement("p", null, "For providing us with the fundamental financial support needed in order to create and fulfill Conjugaid, and for providing us with laboratory facilities. "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://english.ida.dk/", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "IDA"),
React.createElement("p", null, "IDA is Denmark\u2019s largest interest group and trade union with more than 30,000 students in the fields of IT, natural Science and engineering. A membership of IDA gives you access to cheap insurances, academic and social events and plenty of opportunities to build networks."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.pharmadanmark.dk/en", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Pharmadanmark"),
React.createElement("p", null, "Pharmadanmark is a union for professionals and students of life sciences. As specialists in the field, we ensure the best advice for our members both within career and legal counseling. With professional networks and events gives you plenty of opportunities to meet like-minded people within the life science industry."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.youtube.com/user/scishow", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "SciShow"),
React.createElement("p", null, "SciShow is a youtube channel that discusses science news and history and concepts. With equal parts skepticism and enthusiasm, they go a little deeper without going off the deep end. Most of the time, anyway."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.youtube.com/user/crashcourse", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Crash Course"),
React.createElement("p", null, "Tons of awesome courses in one awesome channel! Nicole Sweeney teaches you sociology, Carrie Anne Philbin teaches you computer science, Craig Benzine teaches film history, and Mike Rugnetta is teaching mythology."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "www.idtdna.com", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Integrated DNA Technologies"),
React.createElement("p", null, "Integrated DNA Technologies, Inc. (IDT) develops, manufactures, and markets nucleic acid products for the life sciences industry in the areas of academic research, biotechnology, agriculture, medical diagnostics, and pharmaceutical development. The company's primary business is the production of custom oligonucleotides for molecular biology applications."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.snapgene.com/", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Snapgene"),
React.createElement("p", null, "SnapGene offers the fastest, easiest way to plan, visualize, document, and share your everyday molecular biology procedures. The software is used in more than sixty-five countries, in virtually every major research institution, and in most of the largest pharmaceutical and biotech companies worldwide. Free trials are available from the SnapGene website."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.neb.com/", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "New England Biolabs"),
React.createElement("p", null, "New England Biolabs (NEB) is renowned for consistently providing exceptional product quality and unsurpassed technical support. In addition to NEB\u2019s commitment to scientific innovation and customer relations, NEB notably ensures the environmental sustainability of the company\u2019s business practices. For nearly four decades, NEB has been shaping the landscape of bioscience research by discovering, developing and supporting superior research reagents. Our passion is our promise."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.browserstack.com/", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Browserstack"),
React.createElement("p", null, "Browserstack is an online platform used by web developers and mobile app developers to test how their website or app would work on different browsers, operating systems and mobile devices. This makes it a lot easier to develop a functioning website or app that is functional to all of its users. "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.codecademy.com", target: "_blank" },
React.createElement("img", { src: "https://static.igem.org/mediawiki/2019/7/71/T--SDU-Denmark--codecademy-logo-new.svg" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Codecademy"),
React.createElement("p", null, " Codecademy has helped over 45 million people upgrade their careers with engaging, accessible, and flexible education on programming and data skills. We provide over 200 hands-on interactive lessons ranging from Python to R to Javascript and everything in between. Our learners have gone on to start companies, new jobs, and new lives thanks to what they\u2019ve learned with Codecademy."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.visme.co", target: "_blank" },
React.createElement("img", { src: "https://static.igem.org/mediawiki/2019/f/f5/T--SDU-Denmark--visme-logo.svg" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Visme"),
React.createElement("p", null, "Visme is an easy to use app that allows anyone create better visual content.They empower anyone to visualize ideas into professional Presentations & Infographics. Visme gives you easy (and powerful) tools to manage and empower your online presence. Visme is the tool of choice to create better Visual Content."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("a", { href: "https://www.webflow.com", target: "_blank" },
React.createElement("img", { src: "
" }))),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h3", null, "Webflow"),
React.createElement("p", null, "Webflow is an online platform where you can design, develop, and manage completely custom websites, harnessing all the power of HTML, CSS, and JavaScript in a visual canvas. Webflow is the only design tool that lets you create production-level code.")))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation })));
}
} exports.PartnersSponsors = PartnersSponsors; //# sourceMappingURL=partners.js.map } // default/src/components/Content/Lab_Process_And_Parts/Parts/Parts.jsx $fsx.f[175] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); class Parts extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(174);
});
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "parts", className: "overflow-x-auto" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Parts Catalog"),
React.createElement("div", { className: "under-rubrik" }, "BioBricks used throughout Conjugaid"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "Below you will find a complete list of all the parts that have been used throughout the development of the project. ")),
React.createElement("table", { className: "basic_parts" },
React.createElement("caption", null, "Parts"),
React.createElement("thead", null,
React.createElement("tr", null,
React.createElement("th", null, "Name"),
React.createElement("th", null, "Type"),
React.createElement("th", null, "Description"),
React.createElement("th", { className: "no-dis" }, "Designer"),
React.createElement("th", { className: "no-dis" }, "Length (bp)"))),
React.createElement("tbody", null,
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044024", target: "_blank" }, "BBa_K3044024")),
React.createElement("td", null, "Coding"),
React.createElement("td", null,
"Codon optimized Cas9 for expression in ",
React.createElement("span", { className: "italic" }, "E. coli")),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4107")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044001", target: "_blank" }, "BBa_K3044001")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Strong constitutively active promoter (1.0) and GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044002", target: "_blank" }, "BBa_K3044002")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Strong constitutively active promoter (0.33) and GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044003", target: "_blank" }, "BBa_K3044003")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Strong constitutively active promoter (0.1) and GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044004", target: "_blank" }, "BBa_K3044004")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Strong constitutively active promoter (0.24) and GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044005", target: "_blank" }, "BBa_K3044005")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Strong constitutively active promoter (0.72) and GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044006", target: "_blank" }, "BBa_K3044006")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Constitutively active promoter (0.86) with a GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044007", target: "_blank" }, "BBa_K3044007")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "oriT-R combined with FimH"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "1635")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044008", target: "_blank" }, "BBa_K3044008")),
React.createElement("td", null, "Coding"),
React.createElement("td", null,
"Codon optimized dCas9 for expression in ",
React.createElement("span", { className: "italic" }, "E. coli")),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4107")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044009", target: "_blank" }, "BBa_K3044009")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "dCas9 with IPTG inducible promoter"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4276")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044010", target: "_blank" }, "BBa_K3044010")),
React.createElement("td", null, "RNA"),
React.createElement("td", null, "sgRNA targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "138")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044011", target: "_blank" }, "BBa_K3044011")),
React.createElement("td", null, "RNA"),
React.createElement("td", null, "sgRNA targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "138")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044012", target: "_blank" }, "BBa_K3044012")),
React.createElement("td", null, "RNA"),
React.createElement("td", null, "sgRNA targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "138")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044013", target: "_blank" }, "BBa_K3044013")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/dCas9 system targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044014", target: "_blank" }, "BBa_K3044014")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/dCas9 system targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044015", target: "_blank" }, "BBa_K3044015")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/dCas9 system targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044016", target: "_blank" }, "BBa_K3044016")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Conjugative sgRNA/dCas9 system targeting GFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4801")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044017", target: "_blank" }, "BBa_K3044017")),
React.createElement("td", null, "RNA"),
React.createElement("td", null, "sgRNA targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "138")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044018", target: "_blank" }, "BBa_K3044018")),
React.createElement("td", null, "RNA"),
React.createElement("td", null, "sgRNA targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "138")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044019", target: "_blank" }, "BBa_K3044019")),
React.createElement("td", null, "RNA"),
React.createElement("td", null, "sgRNA targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "138")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044020", target: "_blank" }, "BBa_K3044020")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/dCas9 system targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044021", target: "_blank" }, "BBa_K3044021")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/dCas9 system targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044022", target: "_blank" }, "BBa_K3044022")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/dCas9 system targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044023", target: "_blank" }, "BBa_K3044023")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "dCas9 with constitutive active promoter"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4256")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044024", target: "_blank" }, "BBa_K3044024")),
React.createElement("td", null, "Coding"),
React.createElement("td", null,
"Codon optimized Cas9 for expression in ",
React.createElement("span", { className: "italic" }, "E. coli")),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4107")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044025", target: "_blank" }, "BBa_K3044025")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/Cas9 system targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044026", target: "_blank" }, "BBa_K3044026")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/Cas9 system targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044027", target: "_blank" }, "BBa_K3044027")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "sgRNA/Cas9 system targeting mRFP"),
React.createElement("td", { className: "no-dis" }, "Catharina Bang Jensen"),
React.createElement("td", { className: "no-dis" }, "4422")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044028", target: "_blank" }, "BBa_K3044028")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Conjugative dCas9 system"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "4655")),
React.createElement("tr", null,
React.createElement("td", null,
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044029", target: "_blank" }, "BBa_K3044029")),
React.createElement("td", null, "Composite"),
React.createElement("td", null, "Constitutive active promoter (0.86) incl. a GFP coding sequence"),
React.createElement("td", { className: "no-dis" }, "Anne Sofie L\u00F8gstrup Henriksen"),
React.createElement("td", { className: "no-dis" }, "918")))))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Parts = Parts; //# sourceMappingURL=Parts.js.map } // default/src/components/Content/Project/System_Design/Design.jsx $fsx.f[177] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const react_animate_on_scroll_1 = $fsx.r(169); const rodal_1 = $fsx.r(132); const WavesWidget_1 = $fsx.r(110); const topWaves = [
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,96L48,128C96,160,192,224,288,240C384,256,480,224,576,213.3C672,203,768,213,864,229.3C960,245,1056,267,1152,277.3C1248,288,1344,288,1392,288L1440,288L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
}
]; class Design extends React.Component {
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(176);
});
}
constructor(props) {
super(props);
this.state = { visible: false };
window.document.title = `${props.title} | SDU-Denmark`;
}
show() {
document.body.classList.add("rodal-open");
this.setState({ visible: true });
}
hide() {
document.body.classList.remove("rodal-open");
this.setState({ visible: false });
}
render() {
let accordionData = {
backgroundColor: "design-accordion-bg",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "System Design References",
content: `
- <a href="https://aac.asm.org/content/53/6/2227" target="_blank" class="references">Carattoli, A. (2009). Minireview: Resistance Plasmid Families in Enterobacteriaceae. Antimicrobial Agents and Chemotherapy, 53(6), 2227-2238. </a>
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5545573/" target="_blank" class="references">Hoffman, M. et al. (2017). Comparative Sequence Analysis of Multidrug-Resistant IncA/C Plasmids from Salmonella enterica. Front. Microbiol., 8: 1459. </a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/11212334" target="_blank" class="references">Mazel, D. and J. Davies (1999). Antibiotic resistance in microbes. Cell Mol Life Sci. 56(9), 742-754.</a>
- <a href="Huddleston, J. R. (2014). Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes. Dovepress, 7, 167-176. " target="_blank" class="references">Huddleston, J. R. (2014). Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes. Dovepress, 7, 167-176. </a>
- <a href="https://www.annualreviews.org/doi/10.1146/annurev.ge.25.120191.001051" target="_blank" class="references">Mazodier, P. and J. Davies (1991). Gene transfer between distantly related bacteria. Annual Review of Genetics, 25, 147-171. </a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/15636742" target="_blank" class="references">Llosa, M. and F. de la Cruz (2005). Bacterial conjugation: a potential tool for genomic engineering. Res Microbiol, 156(1), 1-6. </a>
- <a href="https://www.sciencedirect.com/science/article/pii/S0168160508003279?via%3Dihub" target="_blank" class="references">Lampkowska, J. et al. (2008). A standardized conjugation protocol to assess antibiotic resistance transfer between lactococcal species. Int J of Food Microbiology, 127(1-2), 172-175. </a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/8950812" target="_blank" class="references">Berg, R. D. (1996). The indigenous gastrointestinal microflora. Trends Microbiol, 4(11), 430-435.</a>
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4343198/" target="_blank" class="references">Hsu, P. D. et al. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278.</a>
- <a href="https://www.researchgate.net/publication/258055787_CRISPR_interference_CRISPRi_for_sequence-specific_control_of_gene_expression" target="_blank" class="references">Larson, M. H. et al. (2013). CRISPR interference (CRISPRi) for sequence-specific control of gene expression. Nature Protocol, 8(11), 2180-2196. </a>
- <a href="" target="_blank" class="references"></a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "design" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Background"),
React.createElement("div", { className: "under-rubrik" }, "Three Steps"),
React.createElement("div", { className: "manchet mb-20" },
React.createElement("p", { className: "" }, "To guide you easily through our project, we would like to present some relevant background information. We have chosen to split this section into three parts:"),
React.createElement("ol", null,
React.createElement("li", null, "Donor and Recipient Design"),
React.createElement("li", null, "Conjugative Delivery"),
React.createElement("li", null, "CRISPR Action")),
React.createElement("p", null, "Keep reading to be taken on a three-step journey explaining the theoretical principles behind our engineered system.")),
React.createElement("img", { id: "", className: "pt-14", src: "" }))),
React.createElement(react_animate_on_scroll_1.default, { animateIn: "fadeIn" },
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: 1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h2", { className: "darker-border" }, "Donor and Recipient Design"),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", null,
"In order to deliver our CRISPR/(d)Cas9-system - remember this is the term we use when referring interchangeably to both the Cas9- and dCas9-systems - to E. coli, we have designed donor bacteria. The phrase donor arises from its ability to transfer genetic information to neighboring bacteria. The donor bacteria have been designed to contain two separate plasmids; one carrying our CRISPR/(d)Cas9-system and one conjugative containing genes required for cell-to-cell transfer of DNA. Thus, the CRISPR/(d)Cas9-system can be efficiently transferred to resistant recipient ",
React.createElement("i", null, "E. coli.")),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", null,
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices", target: "_blank" }, "Associate Professor Ulrik S. Justesen"),
" from Odense University Hospital informed us of the clinical problems associated with oxacillinase-48 (OXA-48), which is a carbapenemase enzyme. It cleaves the class of antibiotics called carbapenems, thereby generating resistance to common antibiotics. The OXA-48 gene is located on a highly transferable plasmid and is often found in E. coli, pathogens of the Enterobacteriaceae family, which are natural residents of the human gut microbiome ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.researchgate.net/publication/298640728_Laboratory_Detection_and_Clinical_Implication_of_Oxacillinase-48_like_Carbapenemase_The_Hidden_Threat", target: "_blank" }, "Yamuna, D. et al. (2016). Laboratory Detection and Clinical Implication of Oxacillinase-48 like Carbapenemase: The Hidden Threat. J. of global infectious diseases, 8, 41-50."))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.sciencedirect.com/science/article/pii/S1021949819300122", target: "_blank" }, "Ding, R-x. et al. (2019). Revisit gut microbiota and its impact on human health and disease. J of Food and Drug Analysis, 3, 623-631."))),
". After receiving valuable inputs from Associate Professor Justesen, we learned that OXA-48 carbapenemase producing bacteria are prevalent world-wide, and ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices", target: "_blank" }, "in Denmark it is especially an issue at our local hospital"),
". Patients carrying OXA-48 resistance are isolated during future hospitalization, leading to restricted human contact. This is mentally straining for the patients, and may cause loneliness and stigmatization ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5330996/", target: "_blank" }, "Tran, K. et al. (2017). The Effect of Hospital Isolation Precautions on Patient Outcomes and Cost of Care: A Multi-Site, Retrospective, Propensity Score-Matched Cohort Study. J Gen Intern Med., 32(3), 262-268."))),
". Furthermore, patients in isolation requires more resources and are a financial burden to the health-care sector ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5330996/", target: "_blank" }, "Tran, K. et al. (2017). The Effect of Hospital Isolation Precautions on Patient Outcomes and Cost of Care: A Multi-Site, Retrospective, Propensity Score-Matched Cohort Study. J Gen Intern Med., 32(3), 262-268."))),
". Bacteria often carry resistance genes on plasmids with a low copy number ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://aac.asm.org/content/53/6/2227", target: "_blank" }, "Carattoli, A. (2009). Minireview: Resistance Plasmid Families in Enterobacteriaceae. Antimicrobial Agents and Chemotherapy, 53(6), 2227-2238. "))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5545573/", target: "_blank" }, "Hoffman, M. et al. (2017). Comparative Sequence Analysis of Multidrug-Resistant IncA/C Plasmids from Salmonella enterica. Front. Microbiol., 8: 1459. "))),
", and this is also the case for genes encoding OXA-48."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", { id: "no-bottom" }, "Continue reading to see how we developed a dynamic conjugative delivery system based on these donor and recipient bacteria.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 }))),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("h2", { className: "whiter-border" }, "Conjugative Delivery"),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", null,
"Genes encoding antibiotic resistance can spread from one bacterium to another through the natural horizontal gene transfer (HGT) process known as conjugation - this is indeed true for genes encoding OXA-48 ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.researchgate.net/publication/298640728_Laboratory_Detection_and_Clinical_Implication_of_Oxacillinase-48_like_Carbapenemase_The_Hidden_Threat", target: "_blank" }, "Yamuna, D. et al. (2016). Laboratory Detection and Clinical Implication of Oxacillinase-48 like Carbapenemase: The Hidden Threat. J. of global infectious diseases, 8, 41-50. "))),
". Conjugation occurs to a great extent in the human intestine, due to the high density of bacteria found here ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.sciencedirect.com/science/article/pii/S1021949819300122", target: "_blank" }, "Ding, R-x. et al. (2019). Revisit gut microbiota and its impact on human health and disease. J of Food and Drug Analysis, 3, 623-631."))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.sciencedirect.com/science/article/pii/S1438422113000167?via%3Dihub&fbclid=IwAR2I23U_mj_zfHGtWmjZdpso8TvCpv2_KWxWvZayRSuHsNd4HoaD0Zxe2Cs", target: "_blank" }, "Carattoli, A. (2013). Plasmids and the spread of resistance. Int. J. of Med. Microbiology, 303(6-7), 298-304. "))),
". This leads to an increased number of resistant bacteria in the human gut microbiome, complicating the medical therapy ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4378521/", target: "_blank" }, "Ventola, C. L. (2015). The Antibiotic Resistance Crisis, Part 1: Causes and Threats. PT, 40(4), 277-283."))),
". Even after successful treatment, patients may still carry the resistant bacteria in their gut microbiome. Thus, the reservoir for spreading antibiotic resistance increases."),
React.createElement("p", null,
"Conjugation is the only method of HGT that involves direct cell-to-cell contact and requires the presence of mobilizing conjugative factors. These factors are often located on plasmids of which the most well-known are R- and F-plasmids, which are self-transmissible (conjugative) and can mediate their own transfer from one bacterium to another. Transfer of the plasmid is initiated at a region called origin of transfer (oriT) where the relaxosome, a protein complex, introduces a nick in one of the DNA strands of the plasmid that is to be transferred. This event is followed by the actual transmission process known as rolling circle replication where the nicked DNA strand is passed through a protein structure termed the sex-pilus, from which it enters the recipient bacteria ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/15636742", target: "_blank" }, "Llosa, M. and F. de la Cruz (2005). Bacterial conjugation: a potential tool for genomic engineering. Res Microbiol, 156(1), 1-6. "))),
". Besides mediating their own transfer, conjugative plasmids can co-transfer non-self-transmissible (non-conjugative) plasmids containing a compatible oriT region. Since no conjugative plasmids are available within the iGEM kit, we assembled our CRISPR/(d)Cas9-system on a plasmid along with an oriT sequence, making it transferable if paired with an appropriate conjugative plasmid."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("p", null, "An advantage of using conjugation as our delivery system is the fact that the recipient bacteria themselves will become new donors, capable of further delivering our CRISPR/(d)Cas9-system to other recipient bacteria. By combining conjugation with our system, our donor bacteria will initiate a chain reaction leading to further dispersal of our CRISPR/(d)Cas9-system. Thereby, populations of E. coli can be sensitized to common antibiotics."),
React.createElement("p", null,
"Many traditional methods for examining the transfer of plasmids by conjugation have relied on the seeding of bacteria on 2D surfaces, such as filters ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.sciencedirect.com/science/article/pii/S0168160508003279?via%3Dihub", target: "_blank" }, "Lampkowska, J. et al. (2008). A standardized conjugation protocol to assess antibiotic resistance transfer between lactococcal species. Int J of Food Microbiology, 127(1-2), 172-175. "))),
". However, we sought to test conjugation in an environment resembling the 3D nature of the human intestine to an extent by performing conjugation experiments in solution. Likewise, we used a high bacterial density in our conjugation experiments, aiming to mimic the high density of bacteria found within the human gastrointestinal tract ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/8950812", target: "_blank" }, "Berg, R. D. (1996). The indigenous gastrointestinal microflora. Trends Microbiol, 4(11), 430-435. "))),
". Keep reading to get insight into our CRISPR/(d)Cas9-system works when delivered to resistant recipient bacteria."))),
React.createElement(react_animate_on_scroll_1.default, { animateIn: "fadeIn" },
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: 1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h2", { className: "darker-border" }, "CRISPR/(d)Cas9 Action "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" })),
React.createElement("h3", null, "CRISPR/(d)Cas9 For Precision Gene Targeting"),
React.createElement("p", null,
"Ever since the discovery of CRISPR (clustered regularly interspaced short palindromic repeats) as a part of bacteria\u2019s natural immune response against bacteriophages, it has been widely used as an efficient and precise tool for genetic editing ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4343198/", target: "_blank" }, "Hsu, P. D. et al. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278. "))),
". Different CRISPR/Cas-systems exist, but the most well-known is the one we utilize in our project; the CRISPR/Cas9-system originating from Streptococcus pyogenes. In S. pyogenes it works as a library containing information about bacteriophages having previously infected the cell. Upon infection by a former invader, Cas9 (CRISPR associated protein 9) will perform a double stranded break (DSB) in the invading DNA, in which genes will be knocked out as a result of failed reparation mechanisms. Thus, further infection of the bacterium is prevented ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6466564/", target: "_blank" }, "Loureiro, A. & G. J. da Silva. (2019) CRISPR-Cas: Converting A Bacterial Defence Mechanism into A State-of-the-Art Genetic Manipulation Tool. Antibiotics (Basel), 8(1): 18."))),
". "),
React.createElement("p", null,
"When utilizing CRISPR as a gene editing tool, two components are required; a single-guide RNA-molecule (sgRNA) and a Cas-protein. Cas-proteins can perform very precise DSBs in the target DNA after forming a complex with the sgRNA. The sgRNA is responsible for identifying the cleavage target by interacting with Cas9 and leading it to a specific position in the target DNA by complementary base pairing between itself and the target DNA sequence ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4343198/", target: "_blank" }, "Hsu, P. D. et al. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278. "))),
". In addition to the sgRNA-DNA interaction, the presence of a three nucleotide PAM sequence (5\u2019-NGG-3\u2019) in the target DNA is crucial for Cas9\u2019s recognition of the target. After binding to the target DNA and recognition of the PAM sequence, Cas9 induces a DSB in the target DNA, which disrupts the function of the gene ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.annualreviews.org/doi/full/10.1146/annurev-biochem-060815-014607?url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org&rfr_dat=cr_pub%3Dpubmed", target: "_blank" }, "Wang, H. et al. (2016). CRISPR/Cas9 in Genome Editing and Beyond. Annual Review of Biochemistry, 85: 227-264. "))),
". "),
React.createElement("p", null,
"One of the advantages of CRISPR/Cas9 is that the target can be easily modified by exchanging the sgRNA. We utilized this potential through our project, since we constructed several dCas9-systems targeting different genes by exchanging the sgRNA ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.annualreviews.org/doi/full/10.1146/annurev-biochem-060815-014607?url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org&rfr_dat=cr_pub%3Dpubmed", target: "_blank" }, "Wang, H. et al. (2016). CRISPR/Cas9 in Genome Editing and Beyond. Annual Review of Biochemistry, 85: 227-264. "))),
"."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "crispr-fig", className: "", src: "
" })),
React.createElement("p", null,
"Multiple Cas-enzymes exist, and within our project we have chosen to utilize Cas9, which creates a DSB in the DNA. Furthermore, we have used the inactive dead Cas9 (dCas9), which sterically blocks the elongation of transcription as a result of the sgRNA-DNA interaction, thus repressing the expression of the target gene ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4343198/", target: "_blank" }, "Hsu, P. D. et al. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278. "))),
". In dCas9 the catalytic domains are inactive, which prevents the protein from cleaving DNA. In contrast to Cas9, dCas9 induces repression of gene expression ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.annualreviews.org/doi/full/10.1146/annurev-biochem-060815-014607?url_ver=Z39.88-2003&rfr_id=ori%3Arid%3Acrossref.org&rfr_dat=cr_pub%3Dpubmed", target: "_blank" }, "Wang, H. et al. (2016). CRISPR/Cas9 in Genome Editing and Beyond. Annual Review of Biochemistry, 85: 227-264. "))),
". We have primarily worked with dCas9, since we assumed that it might be a gentler solution to repress the gene expression instead of inducing a DSB ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://dx.doi.org/10.1128%2FmBio.00928-13", target: "_blank" }, "Gomaa, A. A. et al. (2014) Programmable Removal of Bacterial Strains by Use of Genome-Targeting CRISPR-Cas Systems. mBio, 5(1), e00928-13. "))),
". However, after talking to ",
React.createElement("a", { target: "_blan", href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices" }, "SNIPR Biome"),
" we became aware that working with Cas9 could be a more efficient and durable solution. For that reason, we ended up testing both systems and their efficiency at downregulating fluorescence genes."),
React.createElement("p", null,
React.createElement("span", { className: "font-bold" }, "Are you interested in learning more about how our system could work clinically?"),
" Click ",
React.createElement("button", { className: "", onClick: this.show.bind(this) }, "here"),
"."),
React.createElement(rodal_1.default, { closeOnEsc: true, customStyles: {
width: "80vw",
height: "80vh"
}, visible: this.state.visible, onClose: this.hide.bind(this) },
React.createElement("div", { className: "rodal-box" },
React.createElement("div", { className: "border-solid border-gray-700 border-b " },
React.createElement("h3", null, "Clinical Potential of Conjugaid")),
React.createElement("p", null, "Our system of engineered donor bacteria could function as an encapsulated drug, aimed at carriers of OXA-48 producing bacteria. Consequently, forced isolation of OXA-48 carriers during future hospitalizations is no longer necessary. Furthermore, patients cleared for OXA-48 cannot transmit this resistance to other patients and people, thus eliminating the reservoir. "),
React.createElement("p", null,
"An orally administered tablet could be designed to release the donor strain in the intestine, from where conjugation would spread our designed CRISPR/(d)Cas9-system to antibiotic resistant ",
React.createElement("i", null, "E. coli."),
" After sensitizing them to specific antibiotics, the formerly resistant E. coli can be eradicated with classic antibiotics if infectious."),
React.createElement("p", null,
"An advantage to conjugation, which is of clinical relevance, is the possibility of targeting a broad range of bacterial strains and furthermore allowing a high carrying capacity of genetic material ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.dovepress.com/horizontal-gene-transfer-in-the-human-gastrointestinal-tract-potential-peer-reviewed-fulltext-article-IDR", target: "_blank" }, "Huddleston, J. R. (2014). Horizontal gene transfer in the human gastrointestinal tract: potential spread of antibiotic resistance genes. Dovepress, 7, 167-176. "))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.annualreviews.org/doi/10.1146/annurev.ge.25.120191.001051", target: "_blank" }, "Mazodier, P. and J. Davies (1991). Gene transfer between distantly related bacteria. Annual Review of Genetics, 25, 147-171. "))),
". This could be relevant in the future for sensitizing strains besides E. coli K12 TOP10."))),
React.createElement("p", { id: "no-bottom" }, "Looking forward to see our results? Hit the \u2018Next\u2019 button and have a look. ")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Design = Design; //# sourceMappingURL=Design.js.map } // default/src/components/Content/Project/Demonstrate/Demonstrate.jsx $fsx.f[179] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const WavesWidget_1 = $fsx.r(110); const topWaves = [
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
}
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,96L48,128C96,160,192,224,288,240C384,256,480,224,576,213.3C672,203,768,213,864,229.3C960,245,1056,267,1152,277.3C1248,288,1344,288,1392,288L1440,288L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
}
]; class Demonstrate extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(178);
});
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "demonstrate" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Demonstration"),
React.createElement("div", { className: "under-rubrik" }, "But does it work?"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" },
" ",
React.createElement("p", null, "From the paragraphs below, it is apparent that our team has successfully accomplished these three points:"),
React.createElement("ol", null,
React.createElement("li", { className: "font-bold" }, "Distribution of plasmids by conjugation increases linearly with time in conditions resembling the human intestine."),
React.createElement("li", { className: "font-bold" },
React.createElement("span", { className: "italic" }, "gfp"),
" can be successfully knocked out by our assembled CRISPR/Cas9-system."),
React.createElement("li", { className: "font-bold" }, "Our assembled CRISPR/dCas9-system can be delivered to recipient E. coli via. conjugation.")),
React.createElement("p", null,
"Taking all of these individual demonstrations into account and combining them, we argue that ",
React.createElement("span", { className: "underline" }, "we have successfully demonstrated the ability of our assembled CRISPR/Cas9-system to be delivered to recipient E. coli in the human intestine via conjugation. "),
" "))))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h2", { className: "darker-border" }, "The Aim of Our Project"),
React.createElement("p", { id: "no-bottom" },
"With Conjugaid, our team set out to aid society in resolving one of the most threatening world problems currently facing humanity; antibiotic resistance ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.nature.com/articles/431899a", target: "_blank" }, "Nathan, C. (2004). Antibiotics at the crossroads. Nature, 431, 899-902."))),
". We wanted to create a sustainable solution by dissociating from the current norm of developing new antimicrobial agents to which bacteria will inevitably develop resistance. According to Associate Professor in Clinical Microbiology Ulrik S. Justesen from Odense University Hospital, bacteria producing the oxacillinase-48 (OXA-48) carbapenemase is a significant problem in our hometown, Odense. By engineering a system capable of eliminating the reservoir for spread of resistant bacteria, we can significantly decrease the amount of antibiotic resistance currently present in society. Thus, our team created an E. coli vehicle which conjugatively delivers a CRISPR/Cas-system able to disrupt specific target genes in recipient ",
React.createElement("i", null, "E. coli."),
" ")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("h2", { className: "whiter-border" }, "How We Made It Work"),
React.createElement("h3", null, "Conjugation"),
React.createElement("p", null,
"We wanted to ensure that our system, if implemented clinically, would reach an adequate amount of recipient bacteria in the human intestine within a limited timeframe. In a high cell density liquid solution under rotation, resembling the natural human gut microbiome to an extent, we examined the transfer rate of plasmids between E. coli over time. From these experiments, a linear increase in dispersal of our plasmids over time ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Results", target: "_blank" }, "was observed"),
". We further optimized our project by",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Model", target: "_blank" }, " modeling"),
" the kinetics of plasmid transfer in conditions resembling the human intestine to an extent. With our three modelled systems, which take certain assumptions into account, we demonstrated the potential of our plasmid system being distributed to all recipient gut bacteria. In a clinical context, this means that all recipients of our engineered system can be sensitized to antibiotics."),
React.createElement("h3", null, "CRISPR/Cas"),
React.createElement("p", null,
"As the sgRNA-component of a CRISPR/Cas-system can be exchanged ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4343198/", target: "_blank" }, "Hsu, P. D. et al. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278. "))),
", we initially set out to demonstrate knockout of the gene gfp (encoding green fluorescent protein, GFP). Assuming that sgRNAs targeting antibiotic resistance genes can be readily created and inserted into a CRISPR/Cas-system, successful proof-of-concept targeting gfp shows that knockout of resistance genes is possible. For our engineered system to be fully demonstrated, the functioning CRISPR/Cas-system needs to be delivered to recipient bacteria through bacterial conjugation. ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Results", target: "_blank" }, "We have successfully demonstrated"),
" that a fully assembled CRISPR/dCas9-system can be delivered from an E. coli vehicle to recipient ",
React.createElement("i", null, "E. coli."),
" This system, however, was not significantly optimal in repressing expression of gfp. It was only after receiving advice from ",
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices" }, "SNIPR Biome"),
" that we changed course and began working with the catalytically active endonuclease Cas9 instead of dCas9. We tested knockout of target genes by CRISPR/Cas9, but have not acquired data on the conjugative delivery using this CRISPR/Cas9-system due to time constraints. However, the only sequential differences between Cas9 and dCas9 are two point mutations ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3922765/", target: "_blank" }, "Larson, M. H. et al. (2013). CRISPR interference (CRISPRi) for sequence-specific control of gene expression. Nat Protoc, 8(11), 2180-2196."))),
". Therefore, we argue that our demonstration of successful conjugative delivery of an assembled CRISPR/dCas9-system shows that similar results can be obtained when testing with a CRISPR/Cas9-system. From ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Results", target: "_blank" }, "results"),
" obtained in fluorescence microscopy and fluorescence-activated cell sorting (FACS) analysis, we have demonstrated that our assembled CRISPR/Cas9-system can successfully knockout expression of gfp in ",
React.createElement("i", null, "E. coli."),
" Thus, we have demonstrated the potential of our system targeting specific antibiotic resistance genes."))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Demonstrate = Demonstrate; //# sourceMappingURL=Demonstrate.js.map } // default/src/components/Content/Project/Conjugation_model/Model.jsx $fsx.f[181] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const WavesWidget_1 = $fsx.r(110); const topWaves = [
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
}
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,96L48,128C96,160,192,224,288,240C384,256,480,224,576,213.3C672,203,768,213,864,229.3C960,245,1056,267,1152,277.3C1248,288,1344,288,1392,288L1440,288L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
}
]; class Model extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(180);
});
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "Conjugation Model References",
content: `
- <a href="" target="_blank" class="references">MANGLER ET LINK TIL DENNE KILDE</a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "model" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Conjugation Model"),
React.createElement("div", { className: "under-rubrik" }, "Kinetics of conjugative plasmid transfer"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, " The future clinical implementation of our project is to efficiently deliver and establish the presence of our CRISPR/(d)Cas9-system in the microbiome, downregulating the expression of resistance genes. However, testing of plasmid transfer in the microbiome of patients provides several major challenges, and is beyond the timescope of our project. Instead, we modelled the kinetics of conjugative plasmid transfer in a bacterial mixture resembling the conditions of the human intestine. As such, our model can be used to somewhat predict long-term plasmid transfer based on acquired experimental data. Furthermore, our model can predict the dosage of donor bacteria that should be administered in order to achieve a sufficient distribution of our CRISPR/(d)Cas9-system within a certain timeframe.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row", id: "no-bottom" },
React.createElement("h2", { id: "no-top", className: "darker-border" }, "System #1"),
React.createElement("p", null,
"Our model is based on a paper by Levin & Rice describing the transfer-kinetics of conjugative and non-conjugative plasmids ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/7002727", target: "_blank" }, "Levin, B. R. and V. A. Rice (1980). The kinetics of transfer of nonconjugative plasmids by mobilizing conjugative factors. Genet Res 35(3), 241-259"))),
". The model is constructed as a series of differential equations tracking the development of distinct bacterial populations based on the law of mass action. "),
React.createElement("p", null, "The five populations in our model include: "),
React.createElement("ol", null,
React.createElement("li", null, "R: Recipients only containing a resistance-conferring plasmid distinguishing them from donor cells."),
React.createElement("li", null, "CN: Donors with both the conjugative and the oriT-compatible non-conjugative plasmids."),
React.createElement("li", null, "TC: Transconjugants having received only the conjugative plasmid."),
React.createElement("li", null, "TN: Transconjugants having received only the non-conjugative plasmid."),
React.createElement("li", null, "TCN: Transconjugants having received both the conjugative and the non-conjugative plasmid. ")),
React.createElement("p", null,
"The progress of the three types of transconjugants is probabilistic and determined by the proportion of all successful matings resulting in the transfer of either only the conjugative plasmid (\u03B1",
React.createElement("sub", null, "C"),
"), only the non-conjugative plasmid (\u03B1",
React.createElement("sub", null, "N"),
") or both plasmids (\u03B1",
React.createElement("sub", null, "CN"),
"). Of the transconjugants, only TNs still function as recipients for the conjugative plasmid, forming TCNs. On the other hand, both TCNs and TCs are unable to function as recipients for future matings, instead functioning as donors. Finally, governs the plasmid transfer rate constant (mL cell",
React.createElement("sup", null, "-1"),
" hr",
React.createElement("sup", null, "-1"),
"). A schematic illustration of the system is shown in ",
React.createElement("span", { className: "font-bold" }, "Figure 1."),
" "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 1.1: Schematic illustration of System #1."),
React.createElement("p", { className: "figure_text" }, "Conjugative transfer of the conjugative and non-conjugative plasmids from CNs to Rs results in the transformation of Rs into three distinct transconjugant populations; TNs, TCNs and TCs. The relative distribution of the transconjugants is determined by the probabilistics of plasmid transfer (\u03B1). TNs are further able to act as recipients for the conjugative plasmid, transforming into TCNs.")),
React.createElement("p", null, "In order to simplify our model, we made the following assumptions:"),
React.createElement("ol", null,
React.createElement("li", null, "All bacteria are at a sufficiently high density such that proliferation can be neglected."),
React.createElement("li", null, "Plasmid loss by segregation occurs due to selective pressure at such a minimal rate that it is neglected."),
React.createElement("li", null, "Conjugation occurs randomly at a frequency proportional to the concentrations of cells containing the conjugative plasmid and the recipients (law of mass action)."),
React.createElement("li", null, "The rate of plasmid transfer is the same for all matings."),
React.createElement("li", null, "Cells carrying the conjugative plasmid cannot serve as recipients for either plasmid."),
React.createElement("li", null,
"A constant proportion of all successful matings between CNs or TCNs and Rs results in transfer of only the conjugative plasmid (\u03B1",
React.createElement("sub", null, "C"),
"), a second constant proportion results in transfer of only the non-conjugative plasmid (\u03B1",
React.createElement("sub", null, "N"),
"), and a third constant proportion results in transfer of both plasmids (\u03B1",
React.createElement("sub", null, "CN"),
"), and the sum of all these equal 1 (\u03B1",
React.createElement("sub", null, "C"),
" + \u03B1",
React.createElement("sub", null, "N"),
" + \u03B1",
React.createElement("sub", null, "CN"),
" = 1)."),
React.createElement("li", null,
"A constant proportion of successful matings between CNs or TCNs and TNs results in transfer of the mobilizing plasmid \u03B1",
React.createElement("sub", null, "C"),
" + \u03B1",
React.createElement("sub", null, "CN"),
")."),
React.createElement("li", null, "The transfer of the plasmids and the transition from recipient cells to new donor cells are instantaneous processes."),
React.createElement("li", null, "The plasmid transfer rate constant is the same across the entire period.")),
React.createElement("p", null, "With these assumptions in mind, we now express the following differential equations for each of the bacterial populations. CNs are not included in the equations, as these do not change over time."),
React.createElement("img", { className: "mx-auto equationlong", src: "
" }),
React.createElement("p", null, "As the number of bacteria is assumed to remain constant over time, the rate constant of plasmid transfer can simply be estimated by the change in total transconjugants, dT, (TN + TC - TCN) divided by the product of the average number of CN and R:"),
React.createElement("img", { id: "small-eq", className: "mx-auto", src: "
" }),
React.createElement("p", null,
"All of the parameters in ",
React.createElement("span", { className: "font-bold" }, "equations (1)-(4)"),
" have been estimated based on the results of conjugation experiments with",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Results", target: "_blank" }, " interrupted mating"),
" at different time points. The experiments were performed using the conjugative plasmid RP4-8, and a non-conjugative plasmid containing an oriT site. For simplification, the non-conjugative plasmid was lacking the CRISPR/(d)Cas9-system under the assumptions that this does not significantly alter the transfer rate of the plasmid. The plasmid transfer rate was obtained by fitting a linear relationship of the total transconjugants over time. However, as these experiments have been performed using simplified experimental setups at a single cell density and donor-to-recipient ratio, the predicted time points do not necessarily reflect the actual time required to propagate the plasmids in the human gut microbiome. These models can therefore not be used as an actual measure of treatment time. However, they may be used to directly compare different systems using similar experimental setups. The acquired values are listed in ",
React.createElement("span", { className: "font-bold" }, "Table 1"),
" below:"),
React.createElement("div", { className: "figure_container" },
React.createElement("p", { className: "figure_name text-blue-800" }, "Table 1:"),
React.createElement("p", { className: "figure_text" },
"Parameter values (\u03B3, \u03B1",
React.createElement("sub", null, "C"),
", \u03B1",
React.createElement("sub", null, "N"),
", \u03B1",
React.createElement("sub", null, "CN"),
", CN and R (t = 0)) based on conjugation experiments with conjugative and a non-conjugative plasmids.")),
React.createElement("table", { className: "basic_parts" },
React.createElement("thead", null,
React.createElement("tr", null,
React.createElement("th", null, "Parameter"),
React.createElement("th", null, "Value"))),
React.createElement("tbody", null,
React.createElement("tr", null,
React.createElement("td", null, "\u03B3\t"),
React.createElement("td", null,
"2.174\u00B710",
React.createElement("sup", null, "-11"),
" (mL cell ",
React.createElement("sup", null, "-1"),
" hr ",
React.createElement("sup", null, "-1"),
")")),
React.createElement("tr", null,
React.createElement("td", null,
"\u03B1",
React.createElement("sub", null, "C"),
"\t"),
React.createElement("td", null, "0.3956")),
React.createElement("tr", null,
React.createElement("td", null,
"\u03B1",
React.createElement("sub", null, "N"),
"\t"),
React.createElement("td", null, "0.3105")),
React.createElement("tr", null,
React.createElement("td", null,
"\u03B1",
React.createElement("sub", null, "CN"),
"\t"),
React.createElement("td", null, "0.2939")),
React.createElement("tr", null,
React.createElement("td", null, "CN"),
React.createElement("td", null,
"2.5\u00B710",
React.createElement("sup", null, "9"),
" (cells mL ",
React.createElement("sup", null, "-1"),
")")),
React.createElement("tr", null,
React.createElement("td", null, "R (t = 0)"),
React.createElement("td", null,
"3.2\u00B710",
React.createElement("sup", null, "9"),
" (cells mL ",
React.createElement("sup", null, "-1"),
")")))),
React.createElement("p", null,
"Our predicted model, illustrated in ",
React.createElement("span", { className: "font-bold" }, "Figure 1.2"),
", shows the change in cell concentration (cells mL",
React.createElement("sup", null, "-1"),
") over time (hours) for the R, TC, TN and TCN populations. As expected, the concentration of R decreases over time as these transform to different transconjugants. The TNs experience an initial increase in concentration while the concentration of Rs remains high, followed by a decrease, as the number of Rs able to turn into TNs diminish and the concentration of TNs functioning as recipients for the conjugative plasmid increases. Both TC and TCN concentrations show an initial rapid increase followed by a fading of the curve as the concentration of R decreases."),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "https://static.igem.org/mediawiki/2019/6/6f/T--SDU-Denmark--system1.svg" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 1.2: Model prediction for System #1"),
React.createElement("p", { className: "figure_text" },
"Predicted changes in the concentration (cells mL",
React.createElement("sup", null, "-1"),
") of Rs, TNs, TCs and TCNs over time (hours).")),
React.createElement("p", { id: "no-bottom" },
"Interestingly, a vast proportion of TCs are formed, almost to the level of the TCN-plateau as seen in ",
React.createElement("span", { className: "font-bold" }, "Figure 1.2."),
" These are unable to function as recipients for the non-conjugative plasmid, due to the aforementioned process of fertility inhibition . With the aim of the project in mind, TCs not being able to function as recipients proves as a major hurdle, as the non-conjugative plasmid containing the CRISPR/(d)Cas9-system is required in order to sensitize recipient bacteria in the gut. This means that a large proportion of the recipient bacteria will never receive the CRISPR/(d)Cas9-system and thereby not become sensitive to antibiotics. Clinically, this will present as a limiting factor when eliminating antibiotic resistance reservoirs.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("h2", { className: "whiter-border" }, "System #2"),
React.createElement("p", null,
"We propose a solution to this problem by constructing a system in which the expressions of the conjugative plasmid genes requires the presence of the non-conjugative plasmid in the same cell. Such a system would allow for TCs to continuously function as recipients for the non-conjugative plasmid, transforming into TCNs. Consequently, this eliminates the ability of TCs to function as donors of the conjugative plasmid, providing one potentially limiting factor of this model. A graphical overview of the modified system is illustrated schematically in ",
React.createElement("span", { className: "font-bold" }, "Figure 2.1.")),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figur 2.1: Schematic illustration of System #2."),
React.createElement("p", { className: "figure_text" }, " Conjugative transfer of the conjugative and non-conjugative plasmids from CNs to Rs results in the transformation of Rs into three distinct transconjugant populations; TNs, TCNs and TCs. The relative distribution of the transconjugants is determined by the probabilistics of plasmid transfer (\u03B1). Both TNs and TCs are further able to act as recipients for the conjugative plasmid, transforming into TCNs.")),
React.createElement("p", null,
"The system can now be described by a modified set of differential equations ",
React.createElement("span", { className: "font-bold" }, "(equations (5)-(8))"),
", accounting for the aforementioned alterations."),
React.createElement("img", { className: "mx-auto equationlong", src: "
" }),
React.createElement("p", null,
"We expect the modified system to show similar outcomes in terms of plasmid transfer rate and probabilistics as the original system, hence, the same parameters as listed in ",
React.createElement("span", null, "Table 1"),
" are used for modelling of the modified system. What is immediately apparent from the model prediction graph in ",
React.createElement("span", null, "Figure 2.2"),
" is the vast change in the distribution of the TCNs and TCs compared to the previous model. Expectedly, the TCs follow a trajectory similar to the TN\u2019s, as the concentration of these cells will now be reflected by both the change in recipient concentration as well as the concentration of TCs that now function as recipients. As both TCs and TNs will unavoidably get exposed to conjugative mating and end up receiving both plasmids, the concentration of TCNs increases continuously, reaching a plateau at the level of the initial concentration of recipient cells as seen in ",
React.createElement("span", null, "Figure 2.2.")),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "\nhttps://static.igem.org/mediawiki/2019/b/bc/T--SDU-Denmark--system2.svg\n" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 2.2: Model prediction for System #2."),
React.createElement("p", { className: "figure_text" },
"Predicted changes in the concentration (cells mL",
React.createElement("sup", null, "-1"),
") of Rs, TNs, TCs and TCNs over time (hours).")),
React.createElement("p", null, "As such, our modified model in System #2 seems to prove our proposed system as an effective solution to one of the major hurdles of the original system (an inefficient delivery of the non-conjugative plasmid to the recipient bacteria). The modified model predicts that all recipient bacteria will eventually end up containing both the non-conjugative CRISPR/(d)Cas9-plasmid along with the conjugative plasmid. In the context of therapeutics, the theoretical outcome of such a system is the sensitizing of all recipient gut bacteria to common antibiotics. "))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row", id: "no-bottom" },
React.createElement("h2", { id: "no-top", className: "darker-border" }, "System #3"),
React.createElement("p", null,
"We propose that it might be more efficient to combine the CRISPR/(d)Cas9-system with the conjugative plasmid, thus requiring only the transfer of a single plasmid containing both systems rather than relying on the transfer of two separate plasmids. Therefore, we decided to model such a system to test our hypothesis. This model is based on the same principles of mass action as the previous models, proving a much simpler system when dealing with the transfer of only a single plasmid. The system consists of Rs, CNs as well as a single population of transconjugants, which is still denoted as TCNs. A schematic overview of the modified system is shown in ",
React.createElement("span", null, "Figure 3.1.")),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "
" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figur 3.1: Schematic illustration of System #3."),
React.createElement("p", { className: "figure_text" }, " Conjugative transfer of the conjugative plasmid from CNs to Rs results in the transformation of Rs into a single transconjugant population; TCNs.")),
React.createElement("p", null, "As only a single transconjugant population is formed from conjugative mating between Rs and CNs, this system does not deal with the same probabilistics as the previous systems. This is due to all successful matings resulting in the transfer of the plasmid containing both the CRISPR/(d)Cas9-system and mobilizing conjugative factors. Based on these assumptions, we now propose the following set of differential equations to describe our modified system:"),
React.createElement("img", { className: "mx-auto equaitonshort", src: "
" }),
React.createElement("p", null,
"As we suspect the plasmid transfer rate constant (\u03B3) to be somewhat similar when dealing with the conjugative transfer of only a single plasmid, and wanted to keep parameters constant for direct comparison, we based this value on the same conjugation experiment as previous systems. We are aware that increasing the plasmid size might negatively impact the plasmid transfer rate. However, as our CRISPR/Cas9-system is relatively small (~5 kb) compared with the size of the RP4-8 plasmid (~60 kb), we choose to neglect this impact. The predicted development of the two distinct bacterial populations is shown in ",
React.createElement("span", { className: "font-black" }, "Figure 3.2"),
", from which we see a symmetrical relationship between the two populations, and the concentration of TCNs increases directly proportional to the decrease in Rs. This is also to be expected by observing ",
React.createElement("span", { className: "font-black" }, "differential equation expressions (9) and (10)"),
", as the expressions of the two populations are directly opposite. "),
React.createElement("div", { className: "figure_container" },
React.createElement("img", { id: "", className: "", src: "\nhttps://static.igem.org/mediawiki/2019/9/90/T--SDU-Denmark--system3.svg\n" }),
React.createElement("p", { className: "figure_name text-blue-800" }, "Figure 3.2: Model prediction for System #3."),
React.createElement("p", { className: "figure_text" },
"Predicted changes in the concentration (cells mL",
React.createElement("sup", null, "-1"),
") of Rs and TCNs over time (hours).")),
React.createElement("p", null,
"The immediate benefit of using conjugative delivery of only a single plasmid is that all recipient bacteria eventually receive the plasmid by conjugation, which is also the case in this system as seen in the model prediction graph ",
React.createElement("span", null, "(Figure 3.2)"),
". Besides overcoming the initial major hurdle of the original system (inefficient delivery of the CRISPR/(d)Cas9-system), this model also seems to improve the predicted time of delivery compared to both of the previous systems. Thus, our modified model in System #3 confirms our proposed solution of combining the entire system, containing both the CRISPR/(d)Cas9-system and conjugative genes, on a single plasmid as a valid option for improving our system and project. Unfortunately, due to incompatibility between the conjugative plasmids and the iGEM BioBrick standards, we were unable to combine our system onto a single plasmid and are thereby prevented from confirming our model. Therefore, we highly encourage future iGEM teams to make an effort to compatibilize conjugative plasmids and integrate these into the parts catalog. We believe that an increased availability of conjugative plasmids will expand the existing options of gene transfer and encourage future teams who wish to utilize conjugation as a method of delivery."),
React.createElement("p", { id: "no-bottom" }, "To learn more about our specific laboratory procedures and the parts we have used in our project, click \u2018Next\u2019.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Model = Model; //# sourceMappingURL=Model.js.map } // default/src/components/Content/Lab_Process_And_Parts/Protocols/Protocols.jsx $fsx.f[183] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const AccordionWidget_1 = $fsx.r(129); const Buttons_1 = $fsx.r(104); class Protocols extends React.Component {
constructor(props) {
super(props);
this.state = { visible: false };
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(182);
});
}
show() {
document.body.classList.add("rodal-open");
this.setState({ visible: true });
}
hide() {
document.body.classList.remove("rodal-open");
this.setState({ visible: false });
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "Protocols and SOPs",
itemTitleColor: "text-blue-800",
items: [
{
title: "Protocols",
content: `
<img src="
"> </img>
<a href="https://static.igem.org/mediawiki/2019/9/94/T--SDU-Denmark--protokol_transform_dcas_to_cas.pdf" target="_blank">Transform dCas to Cas</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/c/c1/T--SDU-Denmark--protokol_design_plasmid_target_bacterium.pdf" target="_blank">Design Plasmid to Target Bacterium</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/1/12/T--SDU-Denmark--protokol_design_plasmid_donor_bacterium.pdf" target="_blank">Design Plasmid to Donor Bacterium</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/7/70/T--SDU-Denmark--protokol_delivery_of_crispr.pdf" target="_blank">Delivery of the CRISPR/dCas9-system</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/b/bc/T--SDU-Denmark--protokol-facs.pdf" target="_blank">Fluorescence microscopy and FACS</a>
},
{
title: "SOPs",
content: `
<img src="
"> </img>
<a href="https://static.igem.org/mediawiki/2019/3/3f/T--SDU-Denmark--SOP1-LA-plates-with-antibiotic.pdf" target="_blank">SOP01 - LA plates with antibiotic</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/a/a6/T--SDU-Denmark--SOP2.pdf" target="_blank">SOP 02 - ON Culture of E.coli</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/5/57/T--SDU-Denmark--SOP3.pdf" target="_blank">SOP 03 - Gel Purification</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/7/7a/T--SDU-Denmark--SOP4.pdf" target="_blank">SOP 04 - Colony PCR with MyTaq</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/1/12/T--SDU-Denmark--SOP5.pdf" target="_blank">SOP 05 - Plasmid Miniprep</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/f/f1/T--SDU-Denmark--SOP6.pdf" target="_blank">SOP 06 - TSB Transformation</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/3/35/T--SDU-Denmark--SOP7.pdf" target="_blank">SOP 07 - Fast Digest</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/c/c0/T--SDU-Denmark--SOP8.pdf" target="_blank">SOP 08 - Conjugation Experiment</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/3/38/T--SDU-Denmark--SOP9.pdf" target="_blank">SOP 09 - Ligation</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/f/fa/T--SDU-Denmark--SOP10.pdf" target="_blank">SOP 10 - Phusion PCR</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/b/b2/T--SDU-Denmark--SOP11.pdf" target="_blank">SOP 11 - Bacterial Freezing Stock</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/8/8a/T--SDU-Denmark--SOP12.pdf" target="_blank">SOP 12 - Making LB and LA Media</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/5/52/T--SDU-Denmark--SOP13.pdf" target="_blank">SOP 13 - Agarose Gel DNA</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/8/83/T--SDU-Denmark--SOP14.pdf" target="_blank">SOP 14 - Table Autoclave</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/3/36/T--SDU-Denmark--SOP15.pdf" target="_blank">SOP 15 - Preparing Primers</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/6/68/T--SDU-Denmark--SOP16.pdf" target="_blank">SOP 16 - Conjugation Time Experiment</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/f/f8/T--SDU-Denmark--SOP18.pdf" target="_blank">SOP 18 - Speedy Vac</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/3/3a/T--SDU-Denmark--SOP19.pdf" target="_blank">SOP 19 - Preparing Eurofins Sequencing Samples</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/d/df/T--SDU-Denmark--SOP20.pdf" target="_blank">SOP 20 - Antibiotic Stock Production</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/e/e0/T--SDU-Denmark--sop21.pdf" target="_blank">SOP 21 - RNA Purification using Phase-Lock
Tubes</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/e/ee/T--SDU-Denmark--sop22.pdf" target="_blank">SOP 22 - cDNa synthesis and Real-Time
Quantitative PCR</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/d/d3/T--SDU-Denmark--sop23.pdf" target="_blank">SOP 23 - Fluorescence Microscopy</a>
<img src=""> </img>
<a href="https://static.igem.org/mediawiki/2019/1/1d/T--SDU-Denmark--SOP24.pdf" target="_blank">SOP 24 - Plasmid Miniprep Without Kit</a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "protocols" },
React.createElement("div", { className: "content mb-24" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Protocols & SOPs"),
React.createElement("div", { className: "under-rubrik" }, "Our Methods"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "In this section we share with you all of our protocols and standard operating procedures used throughout our project. They are available for download as PDFs."))),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionData }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Protocols = Protocols; //# sourceMappingURL=Protocols.js.map } // default/src/components/Content/Lab_Process_And_Parts/Safety/Safety.jsx $fsx.f[185] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const WavesWidget_1 = $fsx.r(110); const react_animate_on_scroll_1 = $fsx.r(169); const topWaves = [
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
}
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,96L48,128C96,160,192,224,288,240C384,256,480,224,576,213.3C672,203,768,213,864,229.3C960,245,1056,267,1152,277.3C1248,288,1344,288,1392,288L1440,288L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
}
]; class Safety extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(184);
});
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "Safety References",
content: `
- <a href="https://apps.who.int/iris/bitstream/handle/10665/312266/9789241515528-eng.pdf?ua=1" target="_blank" class="references">Critically important antimicrobials for human medicine, 6th revision. Geneva: World Health Organization; 2019. Licence: CC BY-NC-SA 3.0 IGO</a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/25380207" target="_blank" class="references">Różańska, D. et al. (2014). The role of Escherichia coli strain Nissle 1917 in the gastro-intestinal diseases. Postepy Hig Med Dosw, 68, 1251-1256. </a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "safety" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Safety"),
React.createElement("div", { className: "under-rubrik" }, "Considerations & Implementations"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "Working in the laboratory every day is rewarding if done safely. Keep reading to learn how we addressed the safety aspects of our project.")))),
React.createElement(react_animate_on_scroll_1.default, { animateIn: "fadeIn" },
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h3", { id: "no-top" }, "General Laboratory Safety"),
React.createElement("p", null, "All our work was performed in a biosafety class 1 laboratory (BSL-1). We received relevant safety training regarding work in GMO laboratories, general laboratory safety and security procedures prior to beginning our work through workshops held by Laboratory Technician Simon Rose. In addition to the workshop, we received written safety guides and video materials about specific higher-risk procedures. We were dressed in lab coats while working, and harmful substances were handled wearing appropriate gloves and in a fume hood. This was especially relevant when working with phenol and chloroform for DNA- and RNA-purification. Additional safety equipment, such as facial screens and gloves, were worn to minimize exposure from UV boards during visualization of DNA-bands in agarose gels. "),
React.createElement("p", null, "Our laboratory is equipped with several types of emergency equipment, including multiple eye wash stations, water showers, fire blankets and extinguishers to ensure an appropriate level of safety. If questions regarding laboratory safety arose during the project, we could always get assistance and support from laboratory technicians."),
React.createElement("h3", null, "Safety Regarding Our Project Design"),
React.createElement("p", null, "Treatment of patients using our solution would include ingestion of our donor bacteria, after which the CRISPR/(d)Cas9-system can be conjugatively delivered to other intestinal bacteria. During our project, we worked on prototype proof-of-concept experiments targeting gpf using a non-pathogenic chassis. This approach was chosen instead of performing the experiments on pathogenic isolates directly from Odense University Hospital, in order to minimize health risks. After achieving proof-of-concept, we assembled a CRISPR/dCas9-system targeting CML resistance genes. This system is yet to be tested . "),
React.createElement("p", null, "Choosing to assemble a system targeting CML was partly based on laboratory convenience, existence of SOPs for its use, its prevalence in backbones in the iGEM kit and its resistance mechanism being well characterized. Furthermore, CML was chosen based on biosafety considerations. We could potentially have worked with clinical pathogenic isolates of OXA-48 producing bacteria from Odense University Hospital. However, to maintain a high degree of safety throughout our work, we chose not to. Knowing that the sgRNA in our system is easily changeable, we rest assured that OXA-48 could possibly be targeted in the future. "),
React.createElement("p", null,
"Our recipient bacteria were designed to imitate an OXA-48 carrying ",
React.createElement("i", null, "E. coli."),
" However, since actual OXA-48 carbapenemase-producing Enterobacteriaceae are resistant to carbapenem, it would not be safe to work with that specific resistance in our laboratory work, since carbapenem antibiotics are on the WHO's list of critically important antibiotics ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.who.int/foodsafety/publications/antimicrobials-sixth/en/", target: "_blank" }, "Critically important antimicrobials for human medicine, 6th revision. Geneva: World Health Organization; 2019. Licence: CC BY-NC-SA 3.0 IGO. "))),
"."),
React.createElement("h3", null, "Safety Considerations Regarding the Potential Future of Our System"),
React.createElement("p", null,
"Questions such as implementation of a kill-switch and environmental risks associated with microbiome therapeutics must be taken into account when it comes to the potential future implementation of our system. Treatment with our system should be controlled by clinicians to prevent excessive use, which would otherwise promote selective pressure of the target bacteria. This is important in order to change the status quo and not contribute to the current increase of antibiotic resistance. A heat sensitive kill-switch like the one made by the ",
React.createElement("a", { href: "https://2017.igem.org/Team:Michigan", target: "_blank" }, "iGEM team from Michigan in 2017"),
" (",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K2301000", target: "_blank" }, "BBa_K2301000"),
" - ",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K2301003", target: "_blank" }, "BBa_K2301003"),
") could be implemented in our system, thereby reducing environmental risks, since donor bacteria would then be killed at temperatures below 37 \u2103. "),
React.createElement("p", null,
"Noteworthy is the possibility of our donor bacteria negatively affecting the human microbiome. Thus, the safety of the donor bacteria is imperative, since it is aimed at diseased patients. Probiotic bacteria such as E. coli Nissle 1917 could be considered as a new chassis. This would be safer for the patients and the microbiome would be less affected ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/25380207", target: "_blank" }, "R\u00F3\u017Ca\u0144ska, D. et al. (2014). The role of Escherichia coli strain Nissle 1917 in the gastro-intestinal diseases. Postepy Hig Med Dosw, 68, 1251-1256. "))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://doi.org/10.1016/j.femsim.2004.10.023", target: "_blank" }, "Westendorf, A. M. et al. (2005). Intestinal immunity of Escherichia coli NISSLE 1917: a safe carrier for therapeutic molecules. FEMS Immunology & Medical Microbiology, 43(3), 373-384. "))),
"."),
React.createElement("p", { id: "no-bottom" },
"These reflections of our potential final product are particularly important within future studies of our project - for us as well as for any other ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Future_Teams", target: "_blank" }, "future iGEM teams"),
". Furthermore, the ethical issues related to our project are highly relevant to consider. You can delve deeper into our considerations regarding the bioethical aspects of Conjugaid in the ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices", target: "_blank" }, "Human Practices"),
" section.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Safety = Safety; //# sourceMappingURL=Safety.js.map } // default/src/components/Content/Lab_Process_And_Parts/Notebook/Notebook.jsx $fsx.f[187] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const AccordionWidget_1 = $fsx.r(129); class Notebook extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(186);
});
}
render() {
let accordionDataMarch = {
backgroundColor: "bg-blue-800",
title: "March",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 12",
content: `
03.26.19 Conjugation & CRISPR
Anne Sofie, Catharina, Annemette, Stine, Thea and Emil.
The whole week we tried to transform different parts into our model organism E. coli K12, TOP10 by following SOP6, though without any luck. Agar plates were also made with appropriate antibiotics by following SOP1. Parts that we have tried to transform:
- BBa_K873003
- BBa_J23100
- BBa_J23101
- BBa_J23111
- BBa_I0500
- BBa_I13507
`
},
{
title: "Week 13",
content: `
03.26.19 Conjugation & CRISPR
Anne Sofie, Catharina, Annemette, Stine, Thea and Emil
We figured out that the preparation of TSB buffer for our transformations was not prepared correctly. We may have forgotten to add DMSO, which makes the cells competent for transformation. Therefore, new TSB buffer was made.
Additionally, we were taught how to make a 1% agarose gel for future electrophoresis experiments and plasmid purification.
03.27.19 Conjugation & CRISPR
Annemette, Stine and Catharina
Transformation of the same parts as in week 12 was done following SOP6.
03.30.19 Conjugation & CRISPR
Stine and Anne Sofie
Transformation of the parts listed below was done:
- BBa_J23111
- BBa_J61100
`
},
{
title: "Week 14",
content: `
`
}
]
};
let accordionDataApril = {
backgroundColor: "bg-blue-800",
title: "April",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 14",
content: `
04.01.19 Conjugation & CRISPR
Anne Sofie
Colonies were observed from transformation experiments made the 03.27.19 and 03.30.19. Additionally, no colonies grew on our negative controls.
04.02.19 Target bacteria design / Fluorescens library
Anne Sofie & Emil
Transformation of the parts listed below was done by using SOP6:
- BBa_J23102
- BBa_I13504
04.03.19 Conjugation & CRISPR
Annemette and Catharina
50% glycerol solution was made for bacteria freeze stock of the different parts.
04.04.19 Conjugation & CRISPR
Catharina, Stine, Annemette and Emil
Colony PCR was made on part BBa_I13504, BBa_J23111 and BBa_J23102 by following SOP4 while a 1% agarose gel was prepared to run the PCR product to check for correct insertion of parts.
04.05.19 Conjugation & CRISPR
Catharina and Stine
Freeze stocks were made on the following parts BBa_I13504, BBa_J23111 and BBa_J23102.
`
},
{
title: "Week 15",
content: `
04.14.19 Target bacteria design / Fluorescens library
Annemette, Thea, Stine and Catharina
Using NucleoSpin Plasmid QuickPure 5.3 kit, part BBa_J23111, BBa_J23102, BBa_J23106, BBa_I13507 and BBa_I13504 was purified. The concentrations of the purified plasmids were determined using a NanoDrop machine.
Part BBa_J23106 and BBa_J23102 were digested using restriction enzyme SpeI and PstI. Part BBa_J23111, BBa_I13507 and BBa_I13504 were digested with XbaI and PstI. SOP7 was followed for this purpose.
50 mg/mL chloramphenicol stock solution was made.
Column purification using NucleoSpin Gel and PCR Clean-Up kit was done on part BBa_J23111.
NucleoSpin Gel extraction kit was used to purify part BBa_J23102, BBa_J23106, BBa_I13507 and BBa_I13504. DNA concentrations were determined using the NanoDrop.
Ligation of the following parts was performed following SOP9:
- BBa_J23111 + BBa_I13504
- BBa_J23111 + BBa_I13507
- BBa_J23106 + BBa_I13504
- BBa_J23106 + BBa_I13507
- BBa_J23102 + BBa_I13504
- BBa_J23102 + BBa_I13507
`
},
{
title: "Week 16",
content: `
04.16.19 Target bacteria design / Fluorescens library
Annemette, Anne Sofie, Stine, Thea, Emil and Catharina
Transformation of the ligated parts was performed following SOP6:
- BBa_J23111 + BBa_I13504
- BBa_J23111 + BBa_I13507
- BBa_J23106 + BBa_I13504
- BBa_J23106 + BBa_I13507
- BBa_J23102 + BBa_I13504
- BBa_J23102 + BBa_I13507
04.17.19 Target bacteria design / Fluorescens library
Annemette, Anne Sofie, Stine, Thea, Emil and Catharina
Colony PCR of the following transformations was performed following SOP4. Colony PCR was also run on undigested BBa_J23106 and BBa_J23102 and a bacteria+water sample as controls:
- BBa_J23111 + BBa_I13504
- BBa_J23111 + BBa_I13507
- BBa_J23106 + BBa_I13504
- BBa_J23106 + BBa_I13507
- BBa_J23102 + BBa_I13504
- BBa_J23102 + BBa_I13507
04.19.19 Target bacteria design / Fluorescens library
Annemette
Overnight culture was made on the new composite parts which were verified by colony PCR yesterday.
04.19.19 Target bacteria design / Fluorescens library
Annemette
Freeze stocks were made on the new parts which have been under construction:
- BBa_J23111 + BBa_I13504
- BBa_J23111 + BBa_I13507
- BBa_J23106 + BBa_I13504
- BBa_J23106 + BBa_I13507
- BBa_J23102 + BBa_I13504
- BBa_J23102 + BBa_I13507
`
}
]
};
let accordionDataMay = {
backgroundColor: "bg-blue-800",
title: "May",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 19",
content: `
05.06.19 Conjugation & CRISPR
Stine
Agar plates with different antibiotics were made. The lab team split into two subgroups: conjugation and CRISPR.
05.09.19 Conjugation
Anne Sofie, Stine and Emil
The first conjugation experiment was run. The conjugations were between the following bacterial combinations by following SOP8:
- BBa_J01003 + BBa_I13504
- BBa_J01003 + BBa_I13507
- BBa_I714031 + BBa_I13504
- BBa_I714031 + BBa_I13507
The conjugating samples were plated on agar plates containing appropriate antibiotics to select for transconjugants. As negative controls, BBa_J01003 and BBa_I714031 was plated on an agar plate containing ampicillin and BBa_I13504 and BBa_I13507 were plated on chloramphenicol.
05.10.19 Conjugation
Stine
There were growth on all plates, unfortunately also a few colonies on the negative controls. Therefore, new agar plates were made where the temperature of the fluid agar medium was cooled down to <50℃.
05.13.19 Conjugation
Anne Sofie
From an O/N culture, a 1000x dilution was made of the following parts in selective LB broth for conjugation:
- BBa_J01003 + cml
- BBa_I13504 + cml
- BBa_J01003 + amp
- BBa_I13507 + amp
SOP8 was followed as for the last conjugation experiment carried out on 05.09.19.
05.14.19 Conjugation
Anne Sophie
The results from the conjugation experiments made on 05.13.19 looked suspicious. Apart from the positive controls, growth was only noticed on one of the combination agar plates (Amp+Cml) where the transconjugants of the combination of BBa_I13507 and BBa_I714031 were to be seen. However, since it was the only plate with growth, the results were not considered useful.
Due to exams the following month, lab work was given a lower priority than previously. The conjugation group began optimizing their protocols.
05.15.2019 and 05.16.2019: CRISPR
Annemette, Catharina and Thea
We started the design of our CRISPR/dCas9 system. We downloaded the DNA sequence for the dCas9 in the iGEM kit and codon optimized it to our chassis E. coli. It was designed without a promoter, since we wanted to test different types and strengths of promoters in our system.
Furthermore, the sgRNA’s targeting rfp and gfp were designed. In addition to the scaffold and the 20 nt DNA-interacting domain, the fragments contained a constitutively active promoter, RBS and terminator sequence.
We ordered the synthesis at Integrated DNA technologies as a part of their free delivery to iGEM teams. We look forward to receiving our DNA and get our work started!
`
},
{
title: "Week 20",
content: `
05.13.19 Conjugation
Anne Sofie
From an O/N culture, a 1000x dilution was made of the following parts in selective LB broth for conjugation:
- BBa_J01003 + cml
- BBa_I13504 + cml
- BBa_J01003 + amp
- BBa_I13507 + amp
SOP8 was followed as for the last conjugation experiment carried out on 05.09.19.
05.14.19 Conjugation
Anne Sophie
The results from the conjugation experiments made on 05.13.19 looked suspicious. Apart from the positive controls, growth was only noticed on one of the combination agar plates (Amp+Cml) where the transconjugants of the combination of BBa_I13507 and BBa_I714031 were to be seen. However, since it was the only plate with growth, the results were not considered useful.
Due to exams the following month, lab work was given a lower priority than previously. The conjugation group began optimizing their protocols.
05.15.2019 and 05.16.2019: CRISPR
Annemette, Catharina and Thea
We started the design of our CRISPR/dCas9 system. We downloaded the DNA sequence for the dCas9 in the iGEM kit and codon optimized it to our chassis E. coli. It was designed without a promoter, since we wanted to test different types and strengths of promoters in our system.
Furthermore, the sgRNA’s targeting rfp and gfp were designed. In addition to the scaffold and the 20 nt DNA-interacting domain, the fragments contained a constitutively active promoter, RBS and terminator sequence.
We ordered the synthesis at Integrated DNA technologies as a part of their free delivery to iGEM teams.
`
}
]
};
let accordionDataJune = {
backgroundColor: "bg-blue-800",
title: "June",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 23",
content: `
06.09.19 Conjugation
Stine
Transformation of the following parts were performed by following SOP6:
- BBa_pSB1K3
- BBa_J61002
- BBa_J23110
- BBa_J23114
- BBa_J23105
- BBa_J23111
`
},
{
title: "Week 24",
content: `
06.11.19 Conjugation
Stine, Annemette, Thea and Catharina.
Transformation of the following parts were performed by following SOP6:
- BBa_J23104
- BBa_R0011
Overnight cultures were made on BBa_J23102, BBa_J01003, BBa_I714031, BBa_pSB1K3, BBa_J61002, BBa_J23110, BBa_J23114 and BBa_J23105.
Freeze stock of BBa_pSB1K3 was made.
06.11.2019 CRISPR
Annemette, Catharina, Stine and Thea.
The following parts were transformed into TOP10:
- BBa_J23104 (constitutively active promoter strength 0.72)
- BBa_R0011 (IPTG inducible promoter)
New LA medium was prepared and a freeze stock containing psB1K3 transformed into TOP10 was made.
06.12.19 Conjugation
Stine
Transformation results from 06.09.19 was great. However, only a few colonies were to be seen on the plates from the transformation experiment performed on 06.11.19. Therefore streak plates were made on the colonies from the transformation experiment on 06.11.19.
Conjugation experiment was performed by following SOP8. The bacteria containing the following parts were used as the donor and acceptor respectively:
- BBa_J23102 and BBa_J01003
- BBa_J23102 and BBa_I714031
06.13.19 Conjugation
Stine
Results from conjugation experiments from 06.12.19 once again looked suspicious. Therefore, colony PCR was run on part BBa_J01003 conjugated with BBa_J23102 and BBa_I714031 conjugated with BBa_J23102 by following SOP4 to check for correct part insert on our donor bacteria.
06.15.19 CRISPR
Annemette, Catharina and Thea.
We had finally received our dCas9 fragments and sgRNAS’s from IDT.
We started the assembly of our dCas9 plasmid. Since we ordered the dCas9
in two fragments, we digested them using SOP 07.
One dCas9 part was digested using Mfe and XbaI, the other with Mfe and PstI.
In both cases, 2.1 buffer was used and the samples were incubated at 37 degrees
for 1 hour and 30 minutes. The fragments were purified using column purification.
Furthermore, we repeated the transformation from 06.11 since it did not succeed. Following parts were transformed:
- BBa_J23104 (constitutively active promoter strength 0.72)
- BBa_R0011 (IPTG inducible promoter)
We performed miniprep of kanamycin backbone (pSB1K3) from an O/N culture using the NucleoSpin Quick Pure kit.
06.16.19 CRISPR
Annemette, Catharina and Thea
We were happy to see that there were colonies on our plates from yesterday.
We performed digest (SOP 07) of the pSB1K3 backbone purified yesterday. It was cut with XbaI and PstI to enable insertion of our dCas9 fragments. The backbone was purified from gel using NucleoSpin PCR and gel clean up.
Using SOP 09, the dCas9 fragments were ligated into our pSB1K3 in different backbone:insert ratios.
`
},
{
title: "Week 25",
content: `
06.18.19 Conjugation
Anne Sofie
We did a lot of research to figure out what was wrong with our experimental setup for the previous conjugations, and we figured out that our model organism E. coli, K12, TOP10 is an F-cell meaning that this bacterial strain is not able to serve as a donor for conjugation. This is due to the lack of a conjugative plasmid which enables bacteria to transfer plasmids between different bacterial strains in a contact-dependent manner. Thus, we were given another bacterial strain named XL1-Blue which is a natural carrier of a so called F’-plasmid. This plasmid contains the essential genes for conjugation.
On that basis, we were able to transform our model organism, E. coli, K12, TOP10, containing part BBa_J01003 into a bacteria capable of conjugating. By following SOP8, a conjugation experiment was performed where the XL1-Blue strain served as our donor bacteria and TOP10 served as our recipient bacteria. The conjuating sample was plated on agar plates containing the appropriate antibiotics to select for TOP10 colonies that have been receiving the F’-plasmid.
06.18.2019 CRISPR
Anne Sofie and Thea
We transformed our ligation of the dCas9 fragments in the pSB1K3 backbone from 06.16 into TOP10 using SOP 06.
06.20.2019 Conjugation
Anne Sofie
The conjugation of the F’-plasmid into the bacteria containing part BBa_J01003 from 06.18.19 succeeded, since we saw colonies on the agar plates containing a combination of antibiotics which selected for bacteria containing the BBa_J01003 and the F’-plasmid. Since E. coli, K12, TOP10 is resistant to streptomycin, the agar plate contained tetracycline (F’-plasmid), chloramphenicol (BBa_J01003), and streptomycin (E. coli, K12, TOP10). Furthermore, the negative controls showed no growth which indicated that the XL1-Blue did not have resistance to chloramphenicol and streptomycin prior to conjugation and the bacteria containing the BBa_J01003 did not have resistance to tetracycline prior to conjugation.
06.20.2019 CRISPR
Annemette and Catharina
Since the transformation of dCas9 in pSB1K3 did not succeed the 06.18, we repeated the transformation of the ligations.
06.21.2019 CRISPR and Target bacteria design / Fluorescens library
Annemette, Catharina and Thea
The ligation of the two dCas9 into the pSB1K3 backbone and its transformation into TOP10 was still not successful .
We performed miniprep on several biobrick parts:
- BBa_I13504 (RBS, gfp, terminator)
- BBa_J23110 (constitutively active promoter strength 0.33)
- BBa_J23114 (constitutively active promoter strength 0.10)
- BBa_J23105 (constitutively active promoter strength 0.24)
- BBa_J23104 (constitutively active promoter strength 0.72)
- BBa_R0011 (IPTG inducible promoter)
We got very low concentrations on the purified plasmid and continued to do so for the next couple of weeks. Important note: Make miniprep of ON cultures! Not bacteria grown to start-exponential phase.
Using SOP 07 we digested J23110, J23114, J23105 with SpeI and PstI and I13504
with XbaI and PstI and incubated them. Since we thought that purifying them
using a column was the right thing to do, we could not use our samples.
Our dCas9 fragments were cut again, one with XbaI and Mfe,
the other one with Mfe and PstI. They were ligated together in different
ratios overnight using SOP 09.
06.22.2019 CRISPR and Target bacteria design / Fluorescens library
Annemette, Catharina and Thea
Following SOP 07 we repeated the digest performed on the 06.21. This time,
we purified the digested plasmids on a gel as we were supposed to (SOP 03).
We ligated different strengths of promoters (BBa_J23110, BBa_J23114, BBa_J23105 and BBa_J23104) with gfp from the BBa_I13504 part using SOP 09.
The BBa_K3044008 in pSB1K3 ligation was transformed into TOP10 using SOP 06.
06.23.2019 Conjugation
Stine
A new conjugation experiment was made by following SOP8. Here, the new donor containing the BBa_J01003 part and the F’-plasmid was mated with an acceptor bacteria containing the BBa_J23102 part with a third resistance (ampicillin), to be able to select for transconjugants that have received both the F’-plasmid and the plasmid containing the oriT.
06.23.2019 Target bacteria design / Fluorescens library
Annemette, Catharina, Stine and Thea
The ligations made the 06.22 (gfp with different promoter strengths) were transformed into TOP10 using SOP 06.
`
},
{
title: "Week 26",
content: `
06.24.2019 Conjugation
Stine
The conjugation experiment showed the best results so far. Colonies were to be seen on the agar plates selecting for transconjugants containing both the F’-plasmid and the parts BBa_J01003 and BBa_BBa_J23102.
Since we have started to see more promising results in the conjugation group, the step will be on improving the used conjugation protocol.
06.24.2019 Target bacteria design / Fluorescens library
Annemette, Catharina, Stine and Thea
Only a few plates showed transformants. We ran PCR and gel electrophoresis on them and confirmed that the backbone had religated and GFP not inserted.
06.24.2019 CRISPR
Annemette, Catharina, Thea
Unfortunately, the transformation of the ligated dCas9 parts was not successful.
06.25.2019 Target bacteria design / Fluorescens library
Annemette, Emma and Thea
Miniprep was made on the following parts from O/N-cultures - and we finally got good concentrations again.
- BBa_I13504 (RBS, gfp, terminator)
- pSB1K3 (Kanamycin backbone)
- BBa_J23110 (constitutively active promoter strength 0.33)
- BBa_J23114 (constitutively active promoter strength 0.10)
- BBa_J23105 (constitutively active promoter strength 0.24)
- BBa_J23104 (constitutively active promoter strength 0.72)
- BBa_R0011 (IPTG inducible promoter)
The GFP part was cut with XbaI and PstI while backbones with promoters were cut with Spe and PstI.
This was done to enable insertion of gfp after different promoters.
After incubation, the different parts were purified on a gel and ligated in different backbone:insert ratios.
06.26.2019 Target bacteria design / Fluorescens library
Annemette, Catharina and Thea
The ligations from yesterday as well as the necessary controls were transformed into the new TOP10 and plated onto plates containing the appropriate antibiotic.
06.27.2019 Target bacteria design / Fluorescens library
Annemette, Catharina and Thea
We finally had transformants on our plates!
`
}
]
};
let accordionDataJuly = {
backgroundColor: "bg-blue-800",
title: "July",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 27",
content: `
07.01.2019 Target bacteria design / Fluorescens library
Annemette
Using SOP 04 and SOP 013, colony PCR and gel electrophoresis was performed on the transformations from 06.26 to confirm the ligations of gfp with different promoters. We were happy to see that they had succeeded.
07.02.19 Conjugation
Anne Sofie
In order to optimize the conjugation protocol we made two main modifications in SOP8.
- Skipping the final 1.5 hours of the conjugation time after the addition of 500 µL LB broth to the conjugating sample
- Instead of growing the cultures to an OD600 = 0.6-0.8, the cultures were grown to an OD600 = 0.3-0.5 instead
The different conjugation experiment was performed as described in SOP08 besides the modifications applied above to the procedure.
07.02.2019 CRISPR
Annemette
We tried to repeat the digest of the dCas9 fragments as well as the kanamycin backbone (pSB1K3) that we wanted to insert them in. Again, pSB1K3 was digested with XbaI and PstI while the dCas9 fragments were digested with XbaI + Mfe and Mfe + PstI respectively. After incubation the different fragments were purified on a gel and ligated in different ratios of backbone and dCas9 fragments overnight.
07.03.19 Conjugation
Anne Sofie and Stine
The conjugation results from 07.02.19 did not look convincing. To obtain knowledge about how fast the bacteria that we are running conjugation experiments on grows, experiments to calculate the generation times of the bacteria containing part BBa_J23102 and another bacteria containing part BBa_J01003 and the conjugating F’-plasmid was determined.
The generation time for the bacteria containing part BBa_J23102 was 27 minutes and the generation time for the bacteria containing both the BBa_J01003 and the F’-plasmid was estimated to be 31 minutes.
<img src="
"/>
Figure 1: The figures both provide data on the growth of two bacteria with different plasmid inserts: 16 refers to the bacteria containing the part BBa_J23102 and 29 refers to the bacteria containing part BBa_J01003 and the F’-plasmid. The left graph depicts the rise in absorbance as a function of time given in minutes. The right graph depicts the logarithm of the absorbance as a function of time given in minutes.
07.03.2019 CRISPR
Annemette and Thea
Using SOP 06 the ligations of dCas9 fragments into psB1K3 were transformed into TOP10.
07.04.2019 CRISPR
Catharina and Thea
We got transformants on our plates, we used SOP 04 and 013 to make colony PCR and gel electrophoresis on the colonies. The gel picture below showed that we had finally ligated our two dCas9 fragments into our pSB1K3 backbone. The biobrick number of the assembled dCas9 fragment is BBa_K3044008.
<img src="
"/>
Figure 2
07.05.2019 CRISPR
Annemette
A freeszestock was made of our BBa_K3044008 in pSB1K3.
We digested the BBa_R0011 part (IPTG inducible promoter) in a pSB1A2 backbone and BBa_K3044008 in pSB1K3 using SOP 07. R0011 was cut with SpeI and PstI and purified on a column using NucleoSpin PCR clean up while BBa_K3044008 in pSB1K3 was cut with XbaI and PstI and purified on gel using SOP 03.
Using SOP 09 the fragments were ligated overnight.
Furthermore, a pSB4A5 plasmid was transformed into TOP10 using SOP06.
07.06.2019 CRISPR
Annemette and Thea
The ligations of BBa_K3044008+BBa_R0011 were transformed into TOP10 using SOP 06. Using SOP10, Phusion PCR was performed on BBa_K3044008 in pSB1K3 with the purpose of inserting promoters of different strengths (F1-F3) in front of dCas9. Three different primer sets were used to insert the promoters. The fragments were purified on a column.
NB: Since we do not use our constructs containing BBa_K3044008 with the F1, F2 and F3 promoters in the testing of our system we did not register them on iGEM registry and will not include gel pictures related to them. In our notebook the constructs are named e.g F1+BBa_K3044008 and are always in a pSB1K3 backbone.
07.07.2019 CRISPR
Annemette and Thea
The three promoters (F1-F3) in front of BBa_K3044008 and BBa_K3044008 (in pSB1K3) were digested using SOP 07 and purified. Afterwards the fragments containing the promoters were ligated with BBa_K3044008 in pSB1K3 using SOP 09. They were transformed into TOP10 using SOP06.
Using SOP04 colony PCR was made on the transformants from yesterday. These were run on a gel using SOP13 and it was revealed that the BBa_K3044008 was successfully inserted after the BBa_R0011 part. The gel looked as expected but we forgot to include a control which will be done another day.
`
},
{
title: "Week 28",
content: `
07.08.2019 CRISPR
Annemette and Thea
The PCR products from yesterday were run on a gel with a control (BBa_K3044008 in pSB1K3). It was now revealed that the BBa_K3044008 part was inserted into the BBa_R0011 part as shown in figure 3. The new biobrick is named BBa_K3044009 and is dCas9 under the control of an IPTG inducible promoter in a pSB1A2 backbone.
<img src="
"/>
Figure 3
Furthermore, colony PCR on the constitutively active promoters in front of dCas9 revealed that one of the promoters (F2) was successfully inserted. The BBa_K3044008 part was sent to sequencing.
07.09.2019 CRISPR
Annemette and Thea
Colony PCR (SOP 04) and gel electrophoresis (SOP 013) was run again on other colonies on the transformations from 07.07 containing the F1 and F3 promoters. Since too much PCR product was loaded onto the gel, it was difficult to tell whether the promoters were inserted. A low copy number backbone (pSB2K3) was transformed into TOP10 using SOP06.
07.10.19 Conjugation
Stine and Emil
Since the optimizations of the conjugation SOP8 did not work as intended, a new replicate on the conjugation between the donor bacteria containing part BBa_J01003 and the F’-plasmid and the target bacteria containing part BBa_J23102 was made.
07.11.2019 Conjugation
Stine and Emil
The results from the experiment made on 07.10.19 showed a sufficient amount of colonies on the different selective plates. However, one of the negative controls contained growth. Colony PCR was run on the transconjugants, the donor andt the target bacteria which showed inconsistent bands, ie. the length of the different parts were not consistent with the bands shown on the gel.
07.11.2019 CRISPR
Annemette, Catharina and Thea
The gel electrophoresis from yesterday was repeated but with a smaller volume PCR product loaded on the gel. The results revealed that the F3 promoter was inserted as well. The ligation of F1+BBa_K3044008 (digested 08.07.19) was performed again using SOP 09.
07.12.2019 CRISPR
Annemette, Catharina and Thea
Transformation was performed on the ligation of F1+BBa_K3044008 using SOP 06.
07.13.2019 CRISPR
Annemette and Catharina
Since there were colonies on our negative control, the transformations were discarded. A new digest of F and BBa_K3044008 (in pSB1K3) using SOP 07 and ligation using SOP 09 was performed.
07.13.2019 Conjugation
Emil
Conjugation experiments using the bacteria containing the BBa_J01003 and the F’-plasmid was conjugated into two different recipient bacteria containing either the BBa_J23102 or the BBa_K3044001. SOP8 was followed.
07.14.2019 Conjugation
Emil and Stine
To be able to see a pattern in the conjugation results, we decided to perform the same conjugation experiments as the previous days using the donor containing the F’-plasmid and the part BBa_J01003 and the target bacteria containing either part BBa_J23102 or BBa_K3044001.
Since we were not able to proof if the F’-plasmid has successfully conjugated into the target bacteria, due to the huge size of the plasmid, we had to rely on the agar plates selecting for the right transconjugants.
07.14.2019 CRISPR
Annemette
The ligations from yesterday containing F1+BBa_K3044008 in pSB1K3 were transformed using SOP 06.
`
},
{
title: "Week 29",
content: `
07.15.2019 CRISPR
Annemette and Catharina
Since there were colonies on the plates, a colony PCR was performed using SOP 04. After running the products on a gel using SOP 013 it was clear that the last promoter (F1) was inserted in front of BBa_K3044008 (in pSB1K3). As mentioned earlier, we do not use these constructs in our final system.
07.16.2019 Conjugation
Emil and Stine
Colony PCR was performed on colonies from conjugation experiments using donors containing part BBa_J01003 with the F’ plasmid and recipients either containing part BBa_J23102 or BBa_J61002. Subsequent gel electrophoresis revealed that all colonies contained oriT, however only those that were selected using only Ampicillin contained RFP or GFP.
07.16.2019 CRISPR
Annemette
Our parts containing BBa_K3044008 with different promoters (BBa_K3044009, F1+BBa_K3044008, F2+BBa_K3044008 and F3+BBa_K3044008) were sent to sequencing. Furthermore, these parts as well as our oligos containing the sgRNA’s were digested (SOP 07) and ligated over night using SOP 09.
07.17.2019 Conjugation
Emil and Stine
New conjugation experiments were performed by using the bacteria containing the F’-plasmid and part BBa_J01003 as donors and now a recipient bacteria containing the kanamycin backbone pSB1K3.
07.17.2019 CRISPR
Annemette and Catharina
The ligations of sgRNA into different BBa_K3044008 parts (BBa_K3044009, F1+BBa_K3044008, F2+BBa_K3044008 and F3+BBa_K3044008) from yesterday were transformed into TOP10 using SOP 06.
07.18.2019 Conjugation
Stine
Once again, the results looked suspicious for unknown reasons. Considerations were made regarding the lack of a washing step when harvesting the cells from the late exponential phase. This was thought of due to the cultivation of the donor and the target bacteria grown in their respective selective medium. Therefore a washing step was introduced after harvesting the bacteria.
07.18.2019 CRISPR
Catharina
There were colonies on our transformants. For that reason, colony PCR (SOP 04) and gel electrophoresis (SOP 013) was performed to check whether the ligation was successful or not. Unfortunately, the gel was not as expected.
07.19.2019 Conjugation
Stine
To check whether the colonies from the conjugation experiment on 07.18.19 contained the right parts, colony PCR was run on the colonies that have grown. However, the gel showed that the transconjugants did not contain the part BBa_J01003 as was expected due to the growth of bacteria on the selective agar plate for transconjugants. Therefore, once again, new agar plates containing appropriate antibiotics were made because we suspected the plates not to be selective for the intended colonies.
07.19.2019 CRISPR
Catharina
Colony PCR was performed on more colonies from the transformation the 07.17 using SOP 04.
07.20.2019 CRISPR
Annemette and Catharina
When the PCR products from yesterday were run on a gel (SOP 013) it was clear that the PCR had not succeeded. For that reason, a new colony PCR was performed using SOP 04. This time the colonies were boiled before added to the PCR tubes.
Unfortunately, the gel electrophoresis revealed that this PCR did not succeed either.
Using SOP 07 we performed the digest of the following parts: BBa_K3044009, F1+BBa_K3044008, F2+BBa_K3044008 and F3+BBa_K3044008, BBa_I13504 (containing gfp) and pSB1K3.
07.21.2019 CRISPR
Annemette
The digested parts from yesterday were purified on either gel or column. Using SOP 09 we ligated cut sgRNAs into psB1K3. Furthermore, gfp from the BBa_I13504 part was inserted after promoters of different strengths. After two hours of ligation, the parts were transformed into TOP10 using SOP 06.
`
},
{
title: "Week 30",
content: `
07.22.19 CRISPR
Annemette and Catharina
There were colonies on many of the transformation plates and for that reason a colony PCR (SOP 04) was performed. After running the PCR products on a gel (SOP 013) it was clear that only one of the ligations had succeeded.
The sgRNA’s were once again ligated with our psB1K3 using SOP 09 but this time the samples ligated over night.
07.23.19 CRISPR
Catharina and Thea
The ligations made yesterday (kanamycin backbone containing different sgRNA’s) were transformed into TOP10 using SOP 06.
O/N cultures of parts containing different promoters in front of GFP were made from PCR streak made yesterday.
07.24.19 CRISPR
Catharina and Thea
Since only a few colonies were present on the transformant plates containing sgRNA’s in kanamycin backbone, they were left for further incubation. Later that day, colony PCR was performed using SOP04 on the plates containing colonies and the PCR products were run on a gel using SOP 013. Since there was too much DNA in the samples, the gel was difficult to analyze. It was decided that a new gel would be loaded with a smaller volume of PCR product the following day.
Miniprep and freeze stocks were made of the O/N cultures containing different promoters in front of GFP.
07.25.19 CRISPR
Thea
The PCR products were run on a gel again. This time only 1 uL sample was loaded. We were happy to see that some of the combinations of sgRNA in kanamycin backbone had succeeded, since the PCR products had the proper length of 500 bp.
<img src="
"/>
Figure 4
The new parts are BBa_K3044017, BBa_K3044018 and BBa_K3044019 in pSB1K3 backbone.
07.26.19 CRISPR
Catharina
The new parts were purified using the NucleoSpin kit. A O/N culture of the pSB1K3 backbone was made, so the rest of the sgRNA’s can be digested and inserted the following day.
07.28.19 CRISPR
Catharina
The sgRNA’s (as oligos) and pSB1K3 were digested using SOP07 and ligated over night using SOP 09.
`
},
{
title: "Week 31",
content: `
07.29.19 CRISPR
Catharina and Thea
The ligations from yesterday were transformed into TOP10 using SOP 06.
Freeze stocks were made of the new parts from the 07.25.
07.30.2019 Conjugation
Anne Sofie and Emil
Conjugation experiments were performed using SOP8 by using BBa_J01003 on a Cml backbone as a donor and recipient cells containing the pSB1K3 backbone to be able to select for transconjugants. Corrections for total cell number was done by measuring the OD600before plating on agar plates containing appropriate antibiotics.
07.30.19 CRISPR
Catharina and Thea
We observed colonies on all our transformation plates. To confirm that the sgRNA’s were inserted into the backbone, colony PCR (SOP 04) was performed. The PCR products were run on a gel using SOP 013. We observed that the sgRNA’s were inserted into our kanamycin backbone as illustrated in figure 5 and 6.
<img src="
"/>
Figure 5
<img src="
"/>
Figure 6
The new parts are BBa_K3044010, BBa_K3044011 and BBa_K3044012 in pSB1K3 backbone.
Furthermore, we did a Phusion PCR using SOP10 to turn dCas9 into Cas9, since only two point mutations differ between them. The gel electrophoresis (SOP 013) revealed that the Phusion PCR did not succeeded.
07.31.2019 Conjugation
Anne Sofie and Emil
The conjugation group found out why their experiments were inconsistent. Since the oriT sequence in the BBa_J01003 is not compatible with the oriT found within the F’-plasmid, the transfer of the non-conjugative plasmid containing the part BBa_J01003, cannot for certain be transferred by conjugation. This might be an explanation why we some days saw few colonies on the plates selecting for transconjugants, and some days without any colonies. The explanation for false-positive colonies observed on previously run colony PCR´s is still a mystery. Perhaps the agar medium has been too hot, potentially inactivating the antibiotics.
Due to the detection of this huge mistake regarding the inattention to the non-compatible oriT sequences between the two plasmids, two plasmids were ordered from Addgene, an RP4-8 and a pRK24 plasmid. Both plasmids were checked to contain the right oriT sequences compatible with the BBa_J01003. Additionally, the oriT-F part BBa_I714030 was ordered from iGEM HQ. The oriT-F sequence were checked to be compatible with the F’-plasmid, that we have done our previous conjugation experiments with.
07.31.19 CRISPR
Catharina and Thea
Using SOP10 the Phusion PCR was repeated to convert dCas9 into Cas9. The gel (SOP 013) run afterwards indicated that the PCR used to correct the two mutations was successful.
Another Phusion PCR was run to assemble the two fragments. The second gel electrophoresis (SOP13) run to check for assembly revealed that it did not succeeded.
O/N cultures of several of the new parts made.
08.01.19 CRISPR
Catharina and Thea
Phusion PCR (SOP10) was performed to assemble Cas9 fragments.
Freeze stocks were made from an ON culture of the new parts. The remaining O/N culture was miniprepped. After purifying the sgRNAs (K3044010, K3044011, K3044012, K3044017, K3044018, K3044019) they were digested with EcoRI and XbaI while the BBa_K304409 part were digested with EcoRI and Spe (SOP07) so the dCas9 fragment could be inserted into our sgRNA backbone. After purifying the sgRNA backbones the concentrations were unfortunately very low. The fragments were ligated ( BBa_K304409 + BBa_K3044010, BBa_K3044011, BBa_K3044012, BBa_K3044017, BBa_K3044018, BBa_K3044019) using SOP 09.
We repeated the second phusion PCR (SOP10) from yesterday to assemble the Cas9 fragments. The gel electrophoresis (SOP13) revealed that it succeeded since the bands had the expected length of 3000 bp as illustrated in figure 7.
<img src="
"/>
Figure 7
08.02.19 CRISPR
Catharina and Thea
The ligations made yesterday were transformed into TOP10 using SOP 06.
Our kanamycin backbone (pSB1K3) and our BBa_K304409 were dried in the SpeedyVac and shipped to Team DTU and Team iGEM Amazonas.
Our Cas9 fragment and BBa_K304408+F1 were digested using SOP07.
08.03.19 CRISPR
Annemette and Catharina
Colony PCR (SOP 04) was run on the transformations. Unfortunately, when the products were run on a gel (SOP 013) it was revealed that the ligation of the sgRNA’s and dCas9 had not succeeded.
The Cas9 fragment was digested and ligated into pSB1K3 using SOP 09. The ligations were transformed into TOP010 using SOP 06.
08.04.19 CRISPR
Catharina
Using SOP 04 colony PCR was made of the transformants and the products were run on a gel using SOP013. The gel revealed that the Cas9 ligation did not succeeded.
`
}
]
};
let accordionDataAugust = {
backgroundColor: "bg-blue-800",
title: "August",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 32",
content: `
08.05.2019 Conjugation
Anne Sofie
While waiting on the ordered conjugative plasmids and parts, data on the F’-plasmid was obtained to get an idea of its conjugation abilities. By following SOP8 the F’-plasmid was conjugated from an XL1-Blue strain into our model bacteria E. coli, TOP10, K12 once again. This donor will now be utilized to study the conjugative abilities of the F’-plasmid between the E. coli, TOP10, K12 strain. Corrections for total cell number was done by measuring the OD600 before plating on agar plates containing appropriate antibiotics.
08.05.19 CRISPR
Catharina
After purifying parts containing sgRNA in pSB1K3 (BBa_K3044010, BBa_K3044011, BBa_K3044012, BBa_K3044017, BBa_K3044018, BBa_K3044019), BBa_K3044019, pSB1K3 and the Cas9 fragment were digested using SOP 07.
The backbone containing the sgRNA’s were digested with EcoRI and XbaI to allow insertion of
dCas9 (cut with EcoRI and SpeI) in front of the sgRNA. The digested parts were purified using
either gel purification or PCR clean up. The products were ligated overnight using SOP 09.
08.06.2019 Conjugation
Anne Sofie
The conjugation results produced on 08.05.19 started to look as expected and the dilutions of the conjugating samples started to look more consistent. Therefore, another replicate was made to test for consistency in the conjugation process.
08.06.19 CRISPR
Catharina
The ligations from yesterday were transformed into TOP10 using SOP06.
The Cas9 fragment and our BBa_K304408+F1 were digested using SOP 07.
08.07.2019 Conjugation
Anne Sofie
The number of colonies on the agar plates selecting for transconjugants looked reasonable when comparing the conjugation results from the 08.05.19 and 08.06.19 suggesting that the conjugation group started to get on the right track at last. We are getting optimistic about our future conjugation experiments again.
08.07.19 CRISPR
Catharina
After running colony PCR (SOP 04) and gel electrophoresis (SOP14) it was revealed that the ligation done the 08.05 had not succeeded.
08.08.19 CRISPR
Catharina
We digested the oligos containing our sgRNA’s as well as BBa_K304409 using SOP 07. After several attempts on inserting the dCas9 in the plasmids containing sgRNA, we thought that it might be easier to insert a smaller fragment (the sgRNA) into the dCas9 plasmid ( BBa_K3044009). This time the oligos (sgRNA) was cut with EcoRI and SpeI while the dCas9 plasmids was cut with EcoRI and XbaI to allow insertion of the sgRNA’s in front of dCas9. After purifying the fragments on a column they were ligated overnight following SOP 09.
Furthermore some BBa_K3044009 was digested so the Cas9 fragment could be inserted instead.
08.09.19 CRISPR
Catharina
The ligations were transformed into TOP10 following SOP06.
08.11.2019 Conjugation
Anne Sofie and Emil
Investigations on how the final target bacteria should look like was initiated today. Since resistance genes in plasmids are often found on low-copy-number plasmids, we sought to find such a plasmid. Therefore, transformation of the backbones pSB4C5 and pSB3C5 was performed. These bacteria, will serve as the recipient bacteria for the upcoming conjugation experiments.
`
},
{
title: "Week 33",
content: `
08.12.2019 Conjugation
Anne Sofie
Conjugative plasmids, RP4-8 and pRK24, was received from Addgene. Streak plates were made of the bacterial stab. Additionally, new agar plates containing appropriate antibiotics for the upcoming days for conjugation was made.
08.12.2019 CRISPR
Catharina and Thea
Since the ligation from the 08.08 did not succeed, the constructs were made again (same assemblies as 08.08). The parts were digested (SOP 07) with the same restriction enzymes as last time, but the ligation (SOP 09) was performed in other backbone:insert ratios. The samples were incubated overnight.
08.13.2019 Conjugation
Anne Sofie
By following SOP8, the RP4-8 and pRK24 was conjugated into separate samples of bacteria each containing part BBa_J01003. These new bacteria, now containing the oriT-R part and either the RP4-8 or the pRK24 plasmid, will serve as the new donors which have to prove the ability of bacteria to deliver a non-conjugative plasmid of our own design.
08.13.2019 CRISPR
Catharina and Thea
A digested Cas9 fragment was inserted into the BBa_K3044009 that was digested the 08.08 following SOP 09 with incubation at room temperature for 2 hours.
Both the Cas9/ BBa_K3044009 ligation as well as the BBa_K3044010/sgRNA ligations were transformed into TOP10 using SOP 06.
08.14.2019 CRISPR
Thea
A digested Cas9 fragment was inserted into the BBa_K3044009 that was digested the 08.08 following SOP 09 with incubation at room temperature for 2 hours.
Both the Cas9/ BBa_K3044009 ligation as well as the BBa_K3044010/sgRNA ligations were transformed into TOP10 using SOP 06.
08.14.2019 Target bacteria design / Fluorescens library
Emil and Anne Sofie
Part BBa_I13504 and BBa_J23102 were purified by using NucleoSpin® Plasmid QuickPure kit. The parts were gel purified using GenElute™ Gel Extraction Kit and ligated using SOP7. This new composite part, BBa_K3044006, was meant to work as our target bacteria.
08.15.2019 CRISPR
Annemette, Catharina and Thea
Since there were no colonies on our transformant plates the constructs had to be made again. This time our oligos (containing sgRNA) were digested instead of the plasmids containing them. In addition to ligating the sgRNAs into the BBa_K3044009, the Cas9 fragment was again inserted into the dCas9 plasmid cut with other restriction enzymes.
08.15.2019 Target bacteria design / Fluorescens library
Emil and Anne Sofie
Part BBa_K3044006 and backbone pSB3C5 was transformed into E. coli, K12, TOP10. These bacteria will serve as the target bacteria that we may choose to test our CRISPR/(d)Cas9 system in.
08.15.2019 Conjugation
Anne Sofie and Emil
The conjugation group sought to investigate how to optimize the conjugation process. By consulting with our supervisors we got interested in utilizing FimH hoping to improve and thereby increase the rate of the transfer of plasmids. Therefore, we purified the part BBa_K1850011, BBa_J01003 and backbone pSB1K3. The following two constructs were made:
- BBa_J01003 inserted in a pSB1K3 backbone
- BBa_J01003 and BBa_K1850011 inserted into a pSB1K3 backbone
08.16.2019 CRISPR
Catharina
Since there were colonies on the plates, a colony PCR (SOP 04) was made. Unfortunately, when running the PCR products on a gel (SOP 14) the bands were not as expected which indicated that the ligation did not succeeded.
08.17.2019 Conjugation
Emil
Colony PCR was run on the following parts made on 08.15.19 to check for correct insertion of plasmids as illustrated in figure 8:
- BBa_K1850011
- BBa_K3044006
- pSB3C5 backbone
<img src="
"/>
Figure 8
08.17.2019 CRISPR
Thea
Miniprep was made on O/N cultures containing dCas9 with different promoters ( BBa_K3044009, BBa_K3044008 + F1/F2/F3).
08.17.2019 Conjugation
Emil
Colony PCR was run on pSB1K3 and BBa_K3044007 to check for correct insertion of plasmid and part.
08.18.2019 CRISPR
Thea
Another digest (SOP 07) and ligation (SOP 09) was performed to make our constructs containing dCas9 ( BBa_K3044010) and sgRNA. After incubating the reactions at 37 degrees for digestion, the enzymes were heat inactivated afterwards. Both the cut oligos and BBa_K3044010 plasmid were purified from a column.
`
},
{
title: "Week 34",
content: `
08.19.2019 Conjugation
Emil, Stine and Anne Sofie
The following donor bacteria were designed by conjugating the ordered plasmids as follows:
- The pRK24 plasmid was conjugated into the bacteria containing the part BBa_J01003 in a pSB1K3 backbone
- The pRK24 plasmid was conjugated into the bacteria containing the part BBa_K3044007 in a pSB1K3 backbone
- The RP4-8 plasmid was conjugated into the bacteria containing the part BBa_J01003 in a pSB1K3 backbone
- The RP4-8 plasmid was conjugated into the bacteria containing the part BBa_K3044007 in a pSB1K3 backbone
08.19.2019 CRISPR
Thea
The digested parts from yesterday (sgRNA and BBa_K3044009) were ligated in different ratios using SOP 09. After incubation at room temperature for two hours the ligations were transformed into TOP10 using SOP 06.
08.20.2019 Conjugation
Emil, Stine and Anne Sofie
Transformation of part BBa_I714030 into E. coli, TOP10, K12 by using SOP6.
08.20.2019 CRISPR
Thea
Since there were colonies on our plates, a colony PCR (SOP 04) and gel electrophoresis (SOP 014) was made to confirm the ligations. Unfortunately, the gel showed that the sgRNA’s were not inserted into BBa_K3044009.
08.20.2019 Target bacteria design / Fluorescens library
Emil, Stine and Anne Sofie
Plasmid purification was done on the following parts which are to be used to design some of the target bacteria:
- Backbone pSB4C5
- Backbone pSB3C5
- BBa_K3044006
- BBa_J23110
- BBa_K3044002
08.21.2019 Target bacteria design / Fluorescens library
Stine
Digest on the following parts was performed by using SOP7:
- Backbone pSB4C5
- Backbone pSB3C5
- BBa_K3044006
- BBa_J23110
- BBa_K3044002
Colony PCR was run on BBa_I714030 was done by following SOP4. Figure 9 depicts the result.
<img src="
"/>
Figure 9
08.22.2019 CRISPR
Thea
Another attempt on insering the sgRNAs into our BBa_K2044009 plasmid was made. This time our sgRNA’s (BBa_K2044010-BBa_K2044012, BBa_K2044017-BBa_K2044019 and other sgRNA in pSB1K3 not further used) were cut out of the backbone with EcoRI and SpeI while the BBa_K2044009 was opened by cutting with EcoRI and XbaI using SOP 07. After purification of the sgRNA and BBa_K2044009, they were ligated for two hours at room temperature using SOP 09. Afterwards our ligations were transformed into TOP10 using SOP 06.
08.22.2019 Conjugation
Stine
The donor cells made on 08.19.19 was made again due to suspicious results from the colony PCR that was run on the new donor cells:
- The pRK24 plasmid was conjugated into the bacteria containing the part BBa_J01003 in a pSB1K3 backbone
- The pRK24 plasmid was conjugated into the bacteria containing the part BBa_K3044007 in a pSB1K3 backbone
- The RP4-8 plasmid was conjugated into the bacteria containing the part BBa_J01003 in a pSB1K3 backbone
- The RP4-8 plasmid was conjugated into the bacteria containing the part BBa_K3044007 in a pSB1K3 backbone
08.23.2019 CRISPR
Annemette and Thea
Unfortunately there were no colonies on our transformant plates except very few, small ones on our transformation of the BBa_K2044009 which was used to test if the cells were competent. Due to the very small amount of cells, it is possible that the cells were not competent. For that reason, a new TSB buffer was made and transformation of the remaining dCas9:sgRNA ligation performed using SOP 06.
08.23.2019 Conjugation
Stine and Anne Sofie
Colony PCR was run on the colonies probably containing both the part BBa_K1850011 and BBa_J01003. The gel proved that the correct insert was performed. SOP4 was used.
<img src="
"/>
Figure 10
We started conjugation experiments, that we will use to characterize the part BBa_J01003. The conjugation experiments were carried out by following SOP8. The first donor bacteria contained the part BBa_J01003 in a pSB1K3 backbone apart from the pRK24 plasmid and the acceptor bacteria containing the part pSB4C5 backbone. The second doner contained the same part BBa_J01003 in a pSB1K3 backbone apart from the RP4-8 plasmid.
08.24.2019 CRISPR
Annemette and Thea
Colony PCR (SOP 04) was run on the few transformants. The gel (SOP 013) indicated that one sgRNA was inserted.
NB: This sgRNA/dCas9 system was not used in our future work for which reason the gel picture is not included.
Another digest (SOP 07) and ligation (SOP 09) was performed on the dCas9 and sgRNAs.
08.24.2019 Conjugation
Anne Sofie and Emil
Two parallel conjugation experiments were set up to be able to compare the effect of the conjugation efficiency when FimH is present in the non-conjugative plasmid or not. SOP8 was followed.
08.25.2019 CRISPR
Thea
Transformation of the ligations from yesterday was performed using SOP 06.
`
},
{
title: "Week 35",
content: `
08.26.2019 Conjugation
Anne Sofie and Emil
Based on the data shown below, the transfer of plasmids did not seem to be improved when FimH is present. According to these results we decided not to do any further experiments with FimH as a part of the conjugation system.
<img src="
"/>
Figure 11: The figure depicts the proportion of recipient bacteria that have either received the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids (TCN).
08.26.2019 CRISPR
Annemette and Thea
Colony PCR (SOP 04) was performed on the transformants. The gel electrophoresis (SOP 013) revealed that one sgRNA was inserted into our dCas9 plasmid.
NB: This sgRNA/dCas9 system was not used in our future work for which reason the gel picture is not included.
08.28.2019 Conjugation
Anne Sofie and Stine
Colony PCR was run on colonies containing the pSB4C5 backbone, or BBa_I714030, or BBa_I714030 including the F’-plasmid (PCR not run for the F’-plasmid due to the length). SOP4 was used.
<img src="
"/>
Figure 12
08.29.2019 Conjugation
Anne Sofie
A replicate of the conjugation between the donor containing the part BBa_I714030 including the F’-plasmid and the acceptor pSB4C5 was carried out by following SOP8.
08.29.2019 CRISPR
Annemette and Thea
Miniprep was done on our plasmids containing sgRNAs in pSB1K3 ((BBa_K2044010-BBa_K2044012, BBa_K2044017-BBa_K2044019)) as well as our BBa_K2044009. Using SOP07 our dCas9 plasmid was cut with EcoRI and XbaI while the sgRNA plasmids were cut with EcoRI and SpeI. After heat inactivation and purification the parts were ligated overnight using SOP09.
08.30.2019 CRISPR
Annemette and Thea
Using SOP06 our ligations were transformed into TOP10.
08.31.2019 Conjugation
Anne Sofie
Colony PCR was run on transconjugants from the 08.29.19 by following SOP4. The results are depicted in figure 13 and 14 below.
<img src="
"/>
Figure 13
<img src="
"/>
Figure 14
08.31.2019 CRISPR
Thea
Since our transformation did not succeed, new overnight cultures of BBa_K2044009 and the sgRNAs (BBa_K2044010-BBa_K2044012, BBa_K2044018- BBa_K2044019 as well as other sgRNA in pSB1K3 not further used).
09.01.2019 Conjugation
Emil and Anne Sofie
Data on the different conjugation experiments carried out for the characterization of the oriT-R (BBa_J01003) and the oriT-F (BBa_K1850011) part was made (figure 15).
<img src="
"/>
Figure 15: The figure depicts the proportion of recipient bacteria that have received either the conjugative plasmid (TC), the non-conjugative plasmid (TN), or both plasmids (TCN). The experiment has been carried out for the three different conjugative plasmids (pRK24, RP4-8, and F’) compatible with their respective oriT-sequence.
Based on the results, the conjugation group decided to produce the future conjugation experiments by using the part BBa_J01003.
09.01.2019 CRISPR
Thea
Miniprep was made of the overnight cultures of sgRNAs in pSB1K3, BBa_K3044009 and BBa_K3044009+F1 and digest was performed using SOP 07 (cut with the same restriction enzymes as 08.29.19). After purification the parts were ligated in different ratios overnight.
`
}
]
};
let accordionDataSeptember = {
backgroundColor: "bg-blue-800",
title: "September",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 36",
content: `
09.03.2019 Conjugation
Anne Sofie and Emil
A conjugation experiment between the donor containing the BBa_J01003 on a pSB1K3 backbone including the RP4-8 plasmid and the acceptor containing an pSB4C5 backbone was performed using SOP8. Additionally, another conjugation experiment was made by using a donor containing the composite part BBa_K3044007 in a pSB1K3 backbone.
09.03.2019 CRISPR
Catharina and Thea
The transformation from yesterday was transformed into TOP10 using SOP06.
09.04.2019 CRISPR
Catharina
There were colonies on our transformants. Colony PCR and gel electrophoresis were made on the transformants using SOP 04 and SOP 013. The gel revealed that of our sgRNA’s were successfully inserted into our dCas9 plasmid (figure 16).
<img src="
"/>
Figure 16
NB: More assemblies worked out (sgRNAs in dCas9+F1) but since these are not further used the gel picture is not included.
The new parts BBa_K3044020 + BBa_K3044021 are composed of sgRNA targeting RFP in a dCas9 part.
09.05.2019 CRISPR
Annemette
The BBa_K608016 part from the kit was transformed into TOP10 using SOP 06.
09.08.2019 CRISPR
Catharina and Thea
Using SOP 09 ligation was performed. psB4C5 (low copy number backbone) or psB3C5 (low-medium copy number backbone) were ligated with GFP and RFP (BBa_J23102, BBa_J23110 and BBa_I13504) over night.
`
},
{
title: "Week 37",
content: `
09.09.2019 Conjugation
Anne Sofie and Emil
The conjugation group sought to develop a conjugation protocol to follow the transfer of plasmids over time. SOP16 has been developed for this purpose. However, the experimental setup has been long in the making. Therefore, SOP16 is the final version of the optimized protocol that we were handed. The following days will be an imperfect version of this optimized SOP. The development of the final SOP was done by making trial and error experiments.
For this experiment the donor bacteria contained the oriT-R part, BBa_J01003, in a pSB1K3 backbone. The acceptor bacteria contained the usual pSB4C5 backbone.
The experiments were carried out in duplicates where the donor contained the part BBa_J01003 on a pSB1K3 backbone apart from from RP4-8 plasmid and the recipient bacteria containing the backbone pSB4C5. A five-hour conjugation experiment was performed where the conjugation sample was interrupted each hour to plate on agar plates containing appropriate antibiotics.
09.09.2019 CRISPR
Catharina and Thea
Ligations containing GFP and RFP in low/medium copy number backbones were transformed into TOP10 using SOP 06.
09.12.2019 CRISPR
Catharina
dCas9 with IPTG inducible promoter containing different sgRNAs (BBa_K3044020 + BBa_K3044021 and sgRNA targeting RFP in BBa_K3044009 + F1) were transformed into pSB4C5 and pSB3C5 (containing RFP reporter). As controls pSB1K3 (kanamycin backbone) as well as sgRNA’s in kanamycin backbone (BBa_K3044017 + BBa_K3044018) were transformed as well.
09.13.2019 Conjugation
Emil
Another conjugation experiment over time were carried out in duplicates where the donor contained the part BBa_J01003 on a pSB1K3 backbone apart from from RP4-8 plasmid and the recipient bacteria containing the backbone pSB4C5. Once again the experiment was run for five hours. However, this time the conjugating sample was allocated into six separate subes for incubation. This was done to avoid interrupting the conjugating bacteria each hour. Results on the time experiment on conjugation performed on 09.13.19 is outlined in the following figure.
<img src="
"/>
Figure 17: The figure depict the proportion of recipient bacteria that have received either the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids (TCN) as a function of time.
09.13.2019 CRISPR
Catharina
There were colonies on the transformants plates from yesterday and the controls were as expected.
09.14.2019 CRISPR
Catharina, Henrik and Thea
Miniprep was performed on over night cultures containing sgRNA in pSB1K3 (BBa_K3044010-BBa_K3044012, BBa_K3044019 and other sgRNAs in pSB1K3 not further used ), BBa_K3044009 and BBa_K3044008+F1.
The parts were digested using SOP 07 and purified - sgRNAs on gel and dCas9 on column.
Afterwards, the sgRNAs were ligated over night with dCas9 (BBa_K3044009) in different ratios using SOP 09.
Colony PCR (SOP 04) were run on the transformant from yesterday (BBa_K3044020 + BBa_K3044021 and sgRNA in BBa_K3044009 + F1 ) transformated into a RFP expressing bacterium). When running the PCR products on a gel (SOP 013) it was revealed that the transformation had succeeded.
Using SOP10 we performed a phusion PCR on BBa_K304409 to correct the mutations in dCas9 and turn it into Cas9. These fragments were purified from a gel. Another phusion PCR (SOP10) was performed to assemble the corrected fragments.
09.15.2019 Conjugation
Anne Sofie
For this conjugation experiment, a modification was made regarding the dilutions we made of the conjugation samples each hour. Instead of only using 1 µL of conjugating sample, we took out all 50 µL of the conjugating sample to get the best representation of the bacterial culture. Once again the experiment was run for five hours.
Results on the time experiment on conjugation performed on 09.09.19 is outlined in the following figures.
<img src="
"/>
Figure 16: The figure depict the proportion of recipient bacteria that have received either the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids (TCN) as a function of time.
<img src="
"/>
Figure 17: The figure depict the proportion of recipient bacteria that have received either the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids (TCN) as a function of time.
09.15.2019 CRISPR
Thea
The ligations from yesterday were transformed into TOP10 using SOP 06.
`
},
{
title: "Week 38",
content: `
09.16.2019 CRISPR
Catharina and Thea
Our Cas9 fragment and the parts containing dCas9+sgRNA (BBa_K3044020+BBa_K3044021) were digested with PacI and SphI using SOP 07. These fragments were ligated in different ratios over night using SOP09.
09.17.2019 CRISPR
Annemette, Catharina and Thea
Colony PCR was performed on the transformants from 09.12 using SOP04. When running the PCR products on a gel using SOP013 it was revealed that almost all the ligations had succeeded.
New parts: BBa_K3044022, BBa_K3044013, BBa_K3044014, BBa_K3044015
NB: More assemblies worked out (sgRNAs in dCas9+F1) but since these are not further used the gel picture is not included.
<img src="
"/>
Figure 18
<img src="
"/>
Figure 19
Furthermore colony PCR was performed on our BBa_K304408+F1 as dCas9+F1+sgRNA (targeting rfp) were transformed into our pSB3C5 and pSB4C5 backbones as controls.
09.18.2019 CRISPR
Annemette and Thea
Since it did not work last time, the Cas9 fragment was ligated into part BBa_K3044020 and BBa_K3044021again.
Fluorescence microscopy was performed on ON cultures containing our BBa_K3044020, BBa_K3044021 and BBa_K3044022 dCas9/sgRNA systems transformed into pSB3C5 and pSB4C5 as well as control samples. Controls ( BBa_K3044017, BBa_K3044018, BBa_K3044019 and pSB1K3) were included.
09.19.2019 Target bacteria design / Fluorescens library
Anne Sofie
Plasmid purification was done on the following parts by using SOP5 or the NucleoSpin® Plasmid QuickPure kit:
- pSB4C5 backbone
- pSB3C5 backbone
- BBa_J23102
- BBa_J23110
- BBa_K3044002
- BBa_K3044006
The parts above were purified from a gel which was done by following SOP3 followed by ligation of the parts in the following order:
- pSB4C5 backbone + BBa_J23102
- pSB4C5 backbone + BBa_J23110
- pSB4C5 backbone + BBa_K3044002
- pSB4C5 backbone + BBa_K3044006
- pSB3C5 backbone + BBa_J23102
- pSB3C5 backbone + BBa_J23110
- pSB3C5 backbone + BBa_K3044002
- pSB3C5 backbone + BBa_K3044006
09.19.2019 CRISPR
Annemette and Thea
Miniprep was performed on following parts (BBa_K3044013, BBa_K3044014, BBa_K3044015, BBa_K3044020, BBa_K3044021, BBa_K3044022) and freeze stocks were made.
Fluorescence microscopy was performed on ON cultures containing different parts transformed into psB4C5.
09.20.2019 CRISPR
Annemette and Catharina
Fluorescence microscopy on the parts from yesterday was carried out.
Following parts were transformed into KG22: BBa_K3044009, BBa_K3044020, BBa_K3044021, BBa_K3044022, Cas9+ BBa_K3044020 and Cas9+BBa_K3044021again.
09.20.2019 Target bacteria design / Fluorescens library
Anne Sofie
Transformation of the ligation products from 09.19.19 was carried out.
09.21.2019 Conjugation
Anne Sofie and Emil
A time experiment in conjugation was performed once again as on 09.15.19 to figure out if the progress of the plasmid dispersal can be more uniform and predictable.
Results on the conjugation experiment from 09.15.19 was outlined (figure 20).
<img src="
"/>
Figure 20: The figure depict the proportion of recipient bacteria that have received either the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids (TCN) as a function of time.
09.21.2019 CRISPR
Annemette and Catharina
Colony PCR was performed on the transformations from yesterday using SOP04. When running the products on a gel (SOP13) it was clear that the transformation of BBa_K3044020 + BBa_K3044021 into KG22 had succeeded.
09.17.2019 CRISPR
Annemette
Colony PCR (SOP04) and gel electrophoresis (SOP013) revealed that the ligation had succeeded. New parts: BBa_K3044002 and BBa_K3044029.
09.22.2019 Conjugation
Anne Sofie and Emil
Another replicate of the a time experiment on conjugation was performed as on 09.15.19.
Results from 09.21.19 is outlined in the figure below.
<img src="
"/>
Figure 21: The figures depict the proportion of recipient bacteria that have received either the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids (TCN) as a function of time.
As can be seen, neither of the figures seem to follow the same progress over time.
09.22.2019 CRISPR
Annemette and Catharina
Miniprep of following parts was performed: BBa_J23102, BBa_J23110, BBa_K3044002 and BBa_K3044029 and freeze stocks were made. The plasmids were transformed into KG22 containing part BBa_K3044020 and BBa_K3044021 using SOP06.
`
},
{
title: "Week 39",
content: `
09.23.2019 CRISPR
Annemette and Catharina
Colony PCR (SOP04) and gel electrophoresis (SOP13) revealed that the transformation had succeeded.
09.23.2019 CRISPR
Annemette and Catharina
A lot of dCas9 (BBa_K3044009, BBa_K3044020, BBa_K3044021 BBa_K3044022 and other cunstructs not registrered since we do not use them in the future ) constructs were transformed into TOP10 bacteria containing BBa_J23102, BBa_J23110, BBa_K3044002 and BBa_K3044029.
09.25.2019 CRISPR
Annemette, Catharina and Thea
The new sgRNAs targeting CML resistance arrived and needed to be inserted into a backbone. Both the sgRNAs and the pSB1K3 backbone were digested with EcoRI and PstI using SOP07. After column purification of the oligos and gel purification of the backbone, the parts were ligated over night using SOP09.
Colony PCR (SOP04) was performed on the transformants from yesterday. The gel electrophoresis (SOP13) revealed that the transformation had succeeded.
Cas9+BBa_K3044021 was transformed into KG22.
09.26.2019 Conjugation
Anne Sofie and Emil
For the time experiment today, rotation was applied while the samples were incubating. Rotation was applied to avoid pellet formation over time. Hopefully this adjustment can contribute to a more linear progress of the dispersal of the plasmids over time. Again, the donor contained the part BBa_J01003 apart from the RP4-8 plasmid. The acceptor bacteria contained the pSB4C5 backbone.
09.26.2019 CRISPR
Annemette, Catharina and Thea
The ligations from yesterday containing CML sgRNAs in pSB1K3 were transformed into TOP10 using SOP06.
Transformation of GFP targeting dCa9 systems (BBa_K3044013, BBa_K3044014, BBa_K3044015) into TOP10 containing part BBa_K3044002 and BBa_K3044029.
Colony PCR on transformations from 09.24 revealed that the transformation had succeeded.
09.27.2019 CRISPR
Catharina
Using SOP04 colony PCR was made on more transformants from the 09.24.
09.28.2019 Conjugation
Emil and Anne Sofie
SOP16 was developed and followed for another duplicate of the time experiment on conjugation as performed on 09.26.19.
09.28.2019 CRISPR
Thea
Since there were colonies on most of our plates containing CML sgRNA in pSB1K3, colony PCR was performed using SOP04. The gel electrophoresis (SOP13) afterwards revealed that the ligations had succeeded (figure 22 and 23).
<img src="
"/>
Figure 22
<img src="https://2019.igem.org/File:T--SDU-Denmark--gel_09.28.2.jpg"/>
Figure 23
`
},
]
};
let accordionDataOctober = {
backgroundColor: "bg-blue-800",
title: "October",
itemTitleColor: "text-blue-800",
items: [
{
title: "Week 40",
content: `
10.01.19 Conjugation & CRISPR
Anne Sofie, Emil, Thea, Catharina and Annemette
The data from the final time experiments was processed and the data have never looked that great.
<img src="
"/>
Figure 24: The figure depicts the proportion of the formation of recipient cells that have received either the non-conjugative plasmid (TN), the conjugative plasmid (TC), or both plasmids as a function of time.
Assembly of the CRISPR-plasmid including the parts BBa_K3044013 and BBa_J01003.
Plasmid purification on part BBa_J01003, BBa_K3044013, and BBa_K3044014 was done by using NucleoSpin® Plasmid QuickPure kit.
The parts were digested by following SOP7 and they were ligated by following SOP9 in the following order:
- BBa_K3044013 + BBa_J01003
- BBa_K3044009 + BBa_J01003
- BBa_K3044010 + BBa_J01003
10.01.2019 CRISPR
Annemette, Catharina and Thea
The CML sgRNAs in pSB1K3 and BBa_K3044009 were digested using SOP07 to allow insertion of the sgRNAs in front of dCas9. After purification the sgRNAs were ligated into the dCas9 backbone for two hours using SOP09. They were transformed into TOP10 using SOP06.
FACS was run on our constructs containing BBa_K3044002 and BBa_K3044020 (GFP acceptor) and BBa_K3044013, BBa_K3044014.
Fluorescence microscopy was done on the constructs containing BBa_J23102, BBa_J23110 and BBa_K3044020 and BBa_K304402..
Colony PCR was run on transformants from the 09.26 and the gel electrophoresis (SOP13) revealed that it had succeeded.
10.02.2019 CRISPR
Thea
Colony PCR (SOP04) was run on the transformants containing dCas9 + sgRNA targeting CML. The gel electrophoresis (SOP13) revealed that the ligations had succeeded (figure 25 and 26).
<img src=""/>
Figure 25
<img src="
"/>
Figure 26
10.03.19 Conjugation & CRISPR
Emil
The ligation product from 10.01.19 was transformed into e. Coli, K12, TOP10.
10.03.2019 CRISPR
Annemette, Catharina and Thea
Fluorescence microscopy was performed on bacteria expressing RFP (BBa_J23102/ BBa_J23110) in combination with dCas9/sgRNA systems (BBa_K3044020, BBa_K3044021 and BBa_K3044022)) and a control (BBa_K3044009). It was also done on bacteria expressing GFP (BBa_K3044002) in combination with different dCas9/sgRNA systems (BBa_K3044013 and BBa_K3044014 in BBa_K3044002.
Digested Cas9 fragment was ligated into different dCas9 parts (BBa_K3044021 and BBa_K3044013) parts using SOP09. These ligations were transformed into TOP10 using SOP06.
10.04.2019 CRISPR
Annemette, Catharina and Thea
Colony PCR (SOP04) on the transformations from yesterday revealed that the ligations of Cas9 into BBa_K3044021 and BBa_K3044013 revealed that it had not succeeded.
10.05.19 Conjugation & CRISPR
Anne Sofie
To create the donor bacteria for an experiment to prove an increase in plasmid dispersal, the RP4-8 plasmid was conjugated into an e. Coli, K12, TOP10. SOP8 was used.
10.05.2019 CRISPR
Annemette, Catharina and Thea
Digested Cas9 fragment and the BBa_K3044021 and BBa_K3044013 parts were purified from gel again. This time the fragments were run a gel for an hour. Using SOP09 the fragments were ligated over night.
10.05.19 Conjugation & CRISPR
Anne Sofie
Plasmid purification was made on BBa_J01003, pSB1K3 backbone, BBa_K3044009, BBa_K3044010, and BBa_K3044013 by following NucleoSpin® Plasmid QuickPure kit.
Digestion was done on the same parts written above by following SOP7 and was further ligated in the following order by using SOP9:
- BBa_K3044013 + BBa_J01003
- BBa_K3044009 + BBa_J01003
- BBa_K3044010 + BBa_J01003
Transformation of the ligation samples written above was done by following SOP6.
10.06.2019 CRISPR
Annemette, Catharina and Thea
The ligations of Cas9 into BBa_K3044021 and BBa_K3044013 were transformed into TOP10 using SOP06. Several sgRNAs (BBa_K3044017, BBa_K3044018, BBa_K3044019) and dCas9/sgRNA systems (BBa_K3044020 and BBa_K3044021 and BBa_K3044009) were transformed into KG22 containing BBa_J23102 using SOP06. Following SOP021 RNA purification of bacteria containing following parts was performed:
- BBa_J23102 (RFP)
- BBa_K3044009 (dCas9) in BBa_J23102
- BBa_K3044021 (dCas9/sgRNA system targeting RFP) in BBa_J23102
- BBa_K3044020 (dCas9/sgRNA system targeting RFP) in BBa_J23102
The quality and concentration of the RNA was checked. This RNA purification is just for practicing for which reason the gel picture is not included.
`
},
{
title: "Week 41",
content: `
10.07.19 Conjugation & CRISPR
Anne Sofie and Emil
Colony PCR was run on the transformation products from 10.06.19 by using SOP7.
<img src="
"/>
Figure 27
<img src="
"/>
Figure 28
10.07.2019 CRISPR
Annemette, Catharina and Thea
Colony PCR on transformations from yesterday revealed that Cas9 was inserted into BBa_K3044021 and BBa_K3044013. The new parts are BBa_K3044027 (Cas9/sgRNA system targeting GFP) and BBa_K3044026 (Cas9/sgRNA system targeting RFP)
Following SOP21 RNA purification was performed on an ON culture of bacteria containing following part combinations:
- BBa_J23102 (RFP with strong promoter)
- BBa_J23110 (RFP with weak promoter)
- BBa_J23102 (RFP with strong promoter) + BBa_K3044009 (dCas9)
- BBa_J23102 (RFP with strong promoter) + BBa_K3044021 (dCas9/sgRNA system targeting RFP)
- BBa_J23102 (RFP with strong promoter) + BBa_K3044020(dCas9/sgRNA system targeting RFP)
- BBa_J23110 (RFP with weak promoter) + BBa_K3044020 (dCas9/sgRNA system targeting RFP)
- BBa_J23110 (RFP with weak promoter) + BBa_K3044025 (Cas9/sgRNA system targeting RFP)
- BBa_J23110 (RFP with weak promoter) + BBa_K3044026 (Cas9/sgRNA system targeting RFP)
- No part (TOP10)
Triplicates were made of all purifications.
<img src="https://2019.igem.org/File:T--SDU-Denmark--gel_10.07.1.jpg"/> <img src="https://2019.igem.org/File:T--SDU-Denmark--gel_10.07.2.jpg"/>
10.08.19 Conjugation & CRISPR
Anne Sofie and Emil
Colony PCR was run on colonies containing part BBa_K3044013 in a pSB1K3 backbone from the transformation performed on 10.06.19 by using SOP4. However, the results did not turn out well.
Furthermore, colony PCR was run on the final conjugation experiment between the donor bacteria containing part BBa_J01003 and the RP4-8 plasmid and the target bacteria containing the pSB4C5 backbone. This gel confirms the presence of the following parts in the respective transconjugants:
- TN: BBa_J01003 and BBa_J04450
- TC: BBa_J04450 (incl. the RP4-8 plasmid which cannot be confirmed by colony PCR and gel electrophoresis. However, the presence of the RP4-8 plasmid was confirmed by selecting for on agar plates containing appropriate antibiotics.
- TNC: BBa_J01003 and BBa_J04450 (incl. the RP4-8 plasmid)
10.08.2019 CRISPR
Annemette, Catharina and Thea
Miniprep made on the new parts. These as well as BBa_K3044010, BBa_K3044012, BBa_K3044013, BBa_K3044014, BBa_K3044015 and BBa_K3044009 are transformed into a KG22 bacterium expressing RFP (BBa_K3044002).
The concentration and quality of the purified RNA from yesterday was measured.
<img src="
"/>
Figure 29
<img src="
"/>
Figure 30
<img src="
"/>
Figure 31
10.09.19 Conjugation & CRISPR
Anne Sofie
Transformation of the newly assembled plasmids, which would serve as the final plasmid constructs for our system was done by following SOP6.
10.09.2019 CRISPR
Annemette, Catharina and Thea
Colony PCR (SOP04) and gel electrophoresis (SOP13) was made on the transformants. Using SOP22 cDNA synthesis and RT-qPCR was made.
10.10.19 Conjugation & CRISPR
Anne Sofie and Emil
Colony PCR was run on the colonies from the transformation experiment performed on 10.09.19 to identify the correct ligated plasmids. One colony from each transformation showed the expected band on the gel.
<img src="
"/>
Figure 32
<img src="
"/>
Figure 33
The RP4-8 plasmid was conjugated into the newly designed bacteria containing the following parts:
- BBa_K3044016
- BBa_K3044028
The conjugation was performed by following SOP8 using the bacteria containing the RP4-8 plasmid as donors and either of the three bacteria containing the parts written above.
10.11.19 Conjugation & CRISPR
Anne Sofie and Emil
Colony PCR was run on the transconjugants from the conjugation experiment on 10.10.19.
Control experiments was made for conjugation to ensure that the transfer of the oriT part (BBa_J01003) is only transferred when the RP4-8 plasmid id present. This was done by designing two other donors; one containing the conjugative RP4-8 plasmid and the pSB1K3 backbone without the oriT-part incorporated and another donor only containing the pSB1K3 backbone with oriT incorporated.
10.12.19 Conjugation & CRISPR
Anne Sofie and Emil
Colony PCR was run on part BBa_K3044016 and BBa_K3044028 due to inconsistent results from 10.11.19.
10.13.19 Conjugation & CRISPR
Anne Sofie
The final experiment to prove the concept of our project was performed by conjugating our final plasmid BBa_K3044023 into the designed accepter bacteria containing the part BBa_K3044002. The experiments were carried out by following SOP8. A control was made as well where the donor contained the part BBa_K3044028 besides the RP4-8 plasmid.
- Donor containing the BBa_K3044016 part and the RP4-8 plasmid and recipient bacteria containing the BBa_K3044002
- Donor containing the BBa_K3044028 part and the RP4-8 plasmid and recipient bacteria containing the BBa_K3044002
10.13.2019 CRISPR
Annemette, Catharina and Thea
Using SOP06 transformation was performed. Following parts: BBa_K3044009, BBa_K3044017, BBa_K3044018, BBa_K3044019, BBa_K3044020, BBa_K3044022, BBa_K3044025, BBa_K3044026 and BBa_K3044027 were transformed into TOP10 expressing RFP (BBa_J23102 + BBa_J23110). Following parts: BBa_K3044009, BBa_K3044010, BBa_K3044011, BBa_K3044012, BBa_K3044015 and BBa_K3044027 were transformed into TOP10 expressing GFP (BBa_K3044002 and BBa_K3044029)
`
},
{
title: "Week 42",
content: `
10.14.2019 CRISPR
Annemette, Catharina and Thea
Using SOP03 colony PCR was performed on the transformants. The transformation had succeeded!
10.15.2019 CRISPR
Annemette, Catharina and Thea
FACS was run on the bacteria containing following constructs:
- No fluorescence (TOP10)
- BBa_K3044002 (GFP with strong promoter)
- BBa_K3044029 (GFP with weak promoter)
- BBa_K3044002 (GFP with strong promoter) + BBa_K3044016 (Conjugative sgRNA-dCas9 system targeting GFP)
- BBa_K3044002 (GFP with strong promoter)+ BBa_K3044028 (Conjugative dCas9 protein)
- BBa_K3044002 (GFP with strong promoter) + 58 BBa_K3044010 (sgRNA targeting GFP)
- BBa_K3044002 (GFP with strong promoter) + BBa_K3044011 (sgRNA targeting GFP)
- BBa_K3044002 (GFP with strong promoter) + BBa_K3044012 (sgRNA targeting GFP)
- BBa_K3044002 (GFP with strong promoter)+ BBa_K3044015 (dCas9/sgRNA system targeting GFP)
- BBa_K3044002 (GFP with strong promoter + BBa_K3044027 (Cas9/sgRNA system targeting GFP)
- BBa_K3044029 (GFP with weak promoter) + BBa_K3044009 (dCas9)
- BBa_K3044029 (GFP with weak promoter) +BBa_K3044010 (sgRNA targeting GFP)
- BBa_K3044029 (GFP with weak promoter) + BBa_K3044011 (sgRNA targeting GFP)
- BBa_K3044029 (GFP with weak promoter) + BBa_K3044012 (sgRNA targeting GFP)
- BBa_K3044029 (GFP with weak promoter) + BBa_K3044015 (dCas9/sgRNA system targeting GFP)
- BBa_K3044029 (GFP with weak promoter) + BBa_K3044027 (Cas9/sgRNA system targeting GFP)
10.15.2019 CRISPR
Annemette, Catharina and Thea
FACS was run on the bacteria containing the same constructs as yesterday.
10.19.19 Conjugation & CRISPR
Anne Sofie
New conjugation experiments were performed with the final plasmid construct, as the experiment performed on 10.13.19. However, a new recipient bacteria containing the part BBa_K3044029 was included. Thereby, four experiments were performed with the following combinations:
- Donor containing the BBa_K3044016 part and the RP4-8 plasmid and recipient bacteria containing the BBa_K3044002
- Donor containing the BBa_K3044028 part and the RP4-8 plasmid and recipient bacteria containing the BBa_K3044002
- Donor containing the BBa_K3044016 part and the RP4-8 plasmid and recipient bacteria containing the BBa_K3044029
- Donor containing the BBa_K3044028 part and the RP4-8 plasmid and recipient bacteria containing the BBa_K3044029
SOP08 was used for this experiment.
10.20.19 Conjugation & CRISPR
Anne Sofie
Colony PCR was made on transconjugants from 10.19.19 according to SOP04. A picture of the gel is shown below:
<img src="
"/>
Figure 34
Colony PCR was made on the final plasmid constructs containing part BBa_K3044016 or part BBa_K3044028. This was done to confirm the presence or absence of the sgRNA, which is the only difference between these parts.
<img src=""/>
Figure 35
`
},
{
title: "Week 43",
content: `
10.21.19 Conjugation & CRISPR
Anne Sofie and Catharina
Facs was run on an overnight culture of the four different transconjugants from 10.19.19 including reference bacteria including the BBa_K3044002 or BBa_K3044029 only.
`
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "notebook" },
React.createElement("div", { className: "content mb-32" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Notebook"),
React.createElement("div", { className: "under-rubrik" }, "Weekly reports"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "Here, we provide detailed reports from each week of our work, explaining what experiments were performed in the laboratory and by which team members."))),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataMarch })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataApril })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataMay })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataJune })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataJuly })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataAugust })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataSeptember })),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataOctober }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Notebook = Notebook; //# sourceMappingURL=Notebook.js.map } // default/src/components/Content/Human_Practices/IHP/IHP.jsx $fsx.f[188] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const WavesWidget_1 = $fsx.r(110); const topWaves = [
{
color: "#BEE1E2",
offset: 0,
path: "M0,96L60,106.7C120,117,240,139,360,170.7C480,203,600,245,720,224C840,203,960,117,1080,85.3C1200,53,1320,75,1380,85.3L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#D0E8E6",
offset: 0,
path: "M0,256L60,229.3C120,203,240,149,360,128C480,107,600,117,720,149.3C840,181,960,235,1080,229.3C1200,224,1320,160,1380,128L1440,96L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
{
color: "#DCEEF2",
offset: 0,
path: "M0,64L60,106.7C120,149,240,235,360,250.7C480,267,600,213,720,165.3C840,117,960,75,1080,85.3C1200,96,1320,160,1380,192L1440,224L1440,320L1380,320C1320,320,1200,320,1080,320C960,320,840,320,720,320C600,320,480,320,360,320C240,320,120,320,60,320L0,320Z"
},
]; const bottomWaves = [
{
color: "#DCEEF2",
offset: 0,
reversed: true,
path: "M0,96L60,85.3C120,75,240,53,360,90.7C480,128,600,224,720,224C840,224,960,128,1080,117.3C1200,107,1320,181,1380,218.7L1440,256L1440,0L1380,0C1320,0,1200,0,1080,0C960,0,840,0,720,0C600,0,480,0,360,0C240,0,120,0,60,0L0,0Z"
},
{
color: "#D0E8E6",
offset: 0,
reversed: true,
path: "M0,64L48,101.3C96,139,192,213,288,218.7C384,224,480,160,576,122.7C672,85,768,75,864,101.3C960,128,1056,192,1152,218.7C1248,245,1344,235,1392,229.3L1440,224L1440,0L1392,0C1344,0,1248,0,1152,0C1056,0,960,0,864,0C768,0,672,0,576,0C480,0,384,0,288,0C192,0,96,0,48,0L0,0Z"
},
{
color: "#BEE1E2",
offset: 0,
reversed: true,
path: "M0,32L34.3,74.7C68.6,117,137,203,206,202.7C274.3,203,343,117,411,101.3C480,85,549,139,617,181.3C685.7,224,754,256,823,266.7C891.4,277,960,267,1029,245.3C1097.1,224,1166,192,1234,197.3C1302.9,203,1371,245,1406,266.7L1440,288L1440,0L1405.7,0C1371.4,0,1303,0,1234,0C1165.7,0,1097,0,1029,0C960,0,891,0,823,0C754.3,0,686,0,617,0C548.6,0,480,0,411,0C342.9,0,274,0,206,0C137.1,0,69,0,34,0L0,0Z"
}
]; class IHP extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "Project & Inspiration References",
content: `
- <a href="" target="_blank" class="references">Filosofisk etik: Normativ etik, praktisk etik og metaetik. Aarhus universitetsforlag, 2011. 2. edition, 2014. Edited by Christensen, A-M. S.</a>
- <a href="https://doi.org/10.1016/j.ijmm.2010.04.003" target="_blank" class="references">Hulscher, M. E. J. L. et al. (2010). Antibiotic use: How to improve it? Int. J. of Med. Microbiology, Vol. 300, Issue 6, 351-356, August, 2010. </a>
- <a href="https://www.nature.com/articles/s41564-018-0327-z" target="_blank" class="references">Peters, J. M. et al. (2019). Enabling genetic analysis of diverse bacteria with Mobile-CRISPRi. Nature Microbiology 4, 244-250. </a>
- <a href="https://apps.who.int/iris/bitstream/handle/10665/312266/9789241515528-eng.pdf?ua=1" target="_blank" class="references">Critically important antimicrobials for human medicine, 6th revision. Geneva: World Health Organization; 2019. Licence: CC BY-NC-SA 3.0 IGO. </a>
- <a href="https://aac.asm.org/content/aac/53/6/2227.full.pdf" target="_blank" class="references">Carattoli, A. (2009). Minireview: Resistance Plasmid Families in Enterobacteriaceae. Antimicrobial Agents and Chemotherapy, 53(6), 2227-2238.</a>
- <a href="https://www.researchgate.net/publication/298640728_Laboratory_Detection_and_Clinical_Implication_of_Oxacillinase-48_like_Carbapenemase_The_Hidden_Threa" target="_blank" class="references">Devi, Y. et al. (2016). Laboratory Detection and Clinical Implication of Oxacilinase-48 like Carbapenemase: The Hidden Threat. Journal of global infectious diseases, 8(1), 41-50. </a>
- <a href="https://dx.doi.org/10.1128%2FmBio.00928-13" target="_blank" class="references">Gomaa, A. A. et al. (2014) Programmable Removal of Bacterial Strains by Use of Genome-Targeting CRISPR-Cas Systems. mBio, 5(1), e00928-13. </a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "IHP" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Human Practices"),
React.createElement("div", { className: "under-rubrik" }, "SDG no. 17: Partnerships for the goals"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "One might have come up with a novel idea and begun pursuing it - but without input from relevant outside parties, this idea will likely never achieve greatness. The creation of innovative solutions requires a two-way dialogue engaging stakeholders potentially implementing them. This way, we can ensure that our project will take into account and integrate anything relevant. Keep reading to learn more about how we have integrated society during the creation of Conjugaid.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: 1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row", id: "no-bottom" },
React.createElement("h2", { className: "darker-border" }, "Bioethics"),
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "darker-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CThe power to control our species\u2019 genetic future is awesome and terrifying. Deciding how to handle it may be the biggest challenge we have ever faced.\u201D"),
React.createElement("p", { className: "quotemaster" }, "- Jennifer A. Doudna, American biochemist and CRISPR-pioneer"),
React.createElement("div", { className: "darker-line" }, " ")),
React.createElement("p", null,
"During our project we had many ethical considerations that we wanted to explore further. Therefore, we talked to MSc student in philosophy Rikke Friis Bentzon, former iGEM team member from ",
React.createElement("a", { href: "https://2016.igem.org/Team:SDU-Denmark", target: "_blank" }, "SDU-Denmark 2016"),
". With a special interest in bioethics, she provided us with the necessary tools needed to investigate the ethics of our project from perspectives."),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null,
"Bioethics is a philosophical branch of ethics that deals with the ethical considerations regarding biotechnology and its methods ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Filosofisk etik: Normativ etik, praktisk etik og metaetik. Aarhus universitetsforlag, 2011. 2. edition, 2014. Edited by Christensen, A-M. S."))),
". Two principles we use to justify Conjugaid is the appeal to well-being and the principle of no harm. Here, the argument is that we should strive to create a happier life ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Filosofisk etik: Normativ etik, praktisk etik og metaetik. Aarhus universitetsforlag, 2011. 2. edition, 2014. Edited by Christensen, A-M. S."))),
". If our system is clinically implemented, it could save the lives of millions of people. Consequently, our project is beneficial in light of the principle of no harm. This principle states that if something is harmless and the user is informed, its use should not be prevented ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Filosofisk etik: Normativ etik, praktisk etik og metaetik. Aarhus universitetsforlag, 2011. 2. edition, 2014. Edited by Christensen, A-M. S."))),
"."),
React.createElement("p", null,
"However, there are arguments contradictory with our project. The first being appeal to naturalness, stating that it is ethically wrong to tamper with nature ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Filosofisk etik: Normativ etik, praktisk etik og metaetik. Aarhus universitetsforlag, 2011. 2. edition, 2014. Edited by Christensen, A-M. S."))),
". When modifying the DNA of an organism, nature is manipulated. When we use synthetic biology to prevent deaths from antibiotic resistant bacterial infections, we are thus affecting the natural process of evolution. The second argument is that we are not yet aware of the consequences of using our donor bacteria in the human gut microbiome. Currently, we do not know if any implications would affect the health of the individual patient, or if we are affecting evolution negatively. The question of potentially affecting the microbiome, however, could be answered by pre-clinical trials on model organisms. "),
React.createElement("p", null,
"The big concern is the long term effects of our system. Will it mutate and create something worse than the antibiotic resistance crisis we are trying to combat? Or might it eventually disrupt the delicate balance in an ecosystem or in nature? These are all questions that we must await future research for answers to. Taking these considerations into account is referred to as the precautionary principle. This principle states that if there is any reason to believe that a practice will cause harm, one should take precautions and progress slowly or not at all ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "", target: "_blank" }, "Filosofisk etik: Normativ etik, praktisk etik og metaetik. Aarhus universitetsforlag, 2011. 2. edition, 2014. Edited by Christensen, A-M. S."))),
". However, the precautionary principle does not always consider the damage potentially caused, if a practise is omitted as a precaution. Perhaps our donor bacteria is harmless and could save the lives of many people. However, we have no way of knowing before the technology is tested. It would therefore be clever to take safety precautions into account when developing our donor bacteria, but we should not be so paralyzed that we do not act with the technology we have at our disposal at this moment. We discussed safety precautions of synthetic biology with Danish legislators during The ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Public_Engagement", target: "_blank" }, "People\u2019s Democratic Festival"),
". Learn more by pressing \u2018play\u2019 on the video below:"),
React.createElement("video", { controls: true },
React.createElement("source", { src: "https://static.igem.org/mediawiki/2019/4/4e/T--SDU-Denmark--fm2.mp4", type: "video/mp4" })),
React.createElement("p", { id: "no-bottom" }, "It might be apparent that we are pro-technology and optimistic about new discoveries. Our belief is that as long as the technology is used on a voluntary and informed basis, it should be allowed. With safety precautions in mind, we could possibly benefit from available technologies to avoid catastrophes such as antibiotic resistance.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row no-bottom" },
React.createElement("h2", { className: "whiter-border" }, "Human Practices"),
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CHear one side and you will be in the dark. Hear both and all will be clear.\u201D"),
React.createElement("p", { className: "quotemaster" }, "- Thomas C. Haliburton, Canadian author."),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null,
"Based on our bioethical considerations and discussions with relevant experts, it was clear to us that increased public information about antibiotic resistance is needed. An issue of this magnitude has to be dealt with by society as a whole just as much as scientists have to work on a solution in the laboratory ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://doi.org/10.1016/j.ijmm.2010.04.003", target: "_blank" }, "Hulscher, M. E. J. L. et al. (2010). Antibiotic use: How to improve it? Int. J. of Med. Microbiology, Vol. 300, Issue 6, 351-356, August, 2010."))),
"."),
React.createElement("p", null, "In April 2019, we were invited by the municipality of Odense to participate in an initiative where 50 young people aged 18-25 could present novel ideas on how our local government can practically implement achieve the SDGs. Our idea about increasing public information on synthetic biology was chosen as one of the five best ideas of the evening, and these \u2018best ideas\u2019 were to be presented to Danish Members of Parliament at The People\u2019s Democratic Festival during the summer. "),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null,
"After attending the event at our municipality, an article featuring one of our team-members discussing the potential of synthetic biology in sustainable development was published on The People\u2019s Democratic Festival\u2019s webpage. ",
React.createElement("a", { href: "https://folkemoedet.dk/nyheder/2019/4/5/engagerede-borgere-skoed-folkehoeringerne-i-gang-i-fredericia-og-odense", target: "_blank" }, "Read it here"),
"."),
React.createElement("p", null,
"During our time at the People\u2019s Democratic Festival (which you can read more about ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Public_Engagement", target: "_blank" }, "here"),
") we learned that synthetic biology is a foreign subject to most Danish politicians. This hinders them from incorporating it in their decision-making, thus depriving the Danish society from potentially valuable technologies. This strengthened our motivation to engage further in increasing public information about this subject."),
React.createElement("p", null, "When performing synthetic biology, it is relevant to carry out risk-assessment. To assess the risk of possible immunological reactions caused by introducing CRISPR in the human gut microbiome, we reached out to Professor Torben Barington from the Department of Clinical Immunology at our local hospital. He pointed out that as a CRISPR/Cas-system is naturally present in bacteria of the gut microbiome, no immunological reactions should take place by introducing our donor bacteria to the patient\u2019s gut microbiome."),
React.createElement("p", null, "Furthermore, we have engaged with MSc student Annemette T\u00F8nnov, currently writing her masters thesis in Political Science. She is debating how the practical use of biotechnology is becoming increasingly available to non-scientists. She interviewed us about working with CRISPR in the laboratory, and the learning-curves associated with DIY-CRISPR."))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: topWaves, top: "", concurrentZDelta: 1 })),
React.createElement("div", { className: "bg-yes" },
React.createElement("div", { className: "content" },
React.createElement("div", { id: "no-bottom", className: "row" },
React.createElement("h2", { className: "darker-border" }, "Integrated Human Practices"),
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "darker-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CComing together is a beginning, staying together is progress, and working together is success.\u201D "),
React.createElement("p", { className: "quotemaster" }, "Henry Ford, American industrialist"),
React.createElement("div", { className: "darker-line" }, " ")),
React.createElement("p", null,
"After being inspired by Peters et al. (2019) ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.nature.com/articles/s41564-018-0327-z", target: "_blank" }, "Peters, J. M. et al. (2019). Enabling genetic analysis of diverse bacteria with Mobile-CRISPRi. Nature Microbiology 4, 244-250. "))),
" and deciding to sensitize bacteria to antibiotics using CRISPR, it was difficult for our team to pinpoint where exactly to begin designing our project. Which resistance genes should we focus on, and which bacteria would be a suitable chassis? To gain answers, we reached out to Ulrik Stenz Justesen, Associate Professor and consultant at the Department of Clinical Microbiology at Odense University Hospital."),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null, "We began our meeting with Justesen by presenting to him our initial project idea. We considered working with urinary tract infections or MRSA (methicillin resistant Staphylococcus aureus). However, Justesen stated the following points as crucial:"),
React.createElement("ol", null,
React.createElement("li", null, "OXA-48 carbepenemase producing Enterobacteriaceae are becoming a critical burden for the Odense University Hospital."),
React.createElement("li", null,
"OXA-48 is prevalent in ",
React.createElement("i", null, "E. coli.")),
React.createElement("li", null,
"The potential target of our system is the human intestine, home to a dense concentration of ",
React.createElement("i", null, "E. coli."))),
React.createElement("p", null, "These points helped shape our project and based on them, we made the following considerations and decisions about what we should be working towards with Conjugaid: "),
React.createElement("ol", null,
React.createElement("li", null,
" ",
React.createElement("p", { className: "underline font-bold" }, "We would aim at targeting OXA-48 resistance."),
React.createElement("p", null,
"After our discussions with Associate Professor Justesen, we soon learned that OXA-48 is a specific resistance often found in pathogenic strains of ",
React.createElement("i", null, "E. coli."),
" OXA-48 is a carbapenemase enzyme, capable of cleaving antibiotics of the carbapenem family. Carbapenem-antibiotics are commonly used to treat infections. Furthermore, they are on WHO\u2019s list of critically important antibiotics ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.who.int/foodsafety/publications/antimicrobials-sixth/en/", target: "_blank" }, "Critically important antimicrobials for human medicine, 6th revision. Geneva: World Health Organization; 2019. Licence: CC BY-NC-SA 3.0 IGO. "))),
". If bacteria develop the ability to produce OXA-48 carbapenemases, they become resistant to carbapenem-antibiotics. Due to safety and accessibility concerns, we chose not to work with OXA-48 carrying bacteria. Instead, we would work on a prototype in which our recipient bacteria is designed to mimic them in the best way possible. Since OXA-48 is located on low-copy number plasmids ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pubmed/19307361", target: "_blank" }, "Carattoli, A. (2009). Minireview: Resistance Plasmid Families in Enterobacteriaceae. Antimicrobial Agents and Chemotherapy, 53(6), 2227-2238."))),
" ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4785756/", target: "_blank" }, "Devi, Y. et al. (2016). Laboratory Detection and Clinical Implication of Oxacilinase-48 like Carbapenemase: The Hidden Threat. Journal of global infectious diseases, 8(1), 41-50. "))),
", our prototype target genes (rfp and gfp) were accordingly expressed on such plasmids. ")),
React.createElement("li", null,
React.createElement("p", { className: "underline font-bold" }, "We would use E. coli as a chassis."),
React.createElement("p", null,
"We learned from Associate Professor Justesen that OXA-48 is prevalent in ",
React.createElement("i", null, "E. coli."),
" Therefore, it made sense to continue to work with E. coli as the chassis in our conjugation experiments.")),
React.createElement("li", null,
React.createElement("p", { className: "underline font-bold" }, "Our experiments should be designed to mimic the environment of the human gastrointestinal tract. "),
React.createElement("p", null, "Since the gastrointestinal tract is a natural environment of E. coli and the place where the spread of OXA-48 occurs (where our system could possibly function), we decided to design our experiments so that they mimic the human intestines. This was achieved by performing the conjugation experiments in solution with a high density of bacteria."))),
React.createElement("p", null, "We integrated the suggestions from Associate Professor Justesen in our project. He also made us consider the potential aspects of our project regarding distribution to patients. Our initial thought was the creation of a \u201Cpre-drug\u201D targeting resistant bacteria in patients being treated with antibiotics. Justesen, however, pointed out that such a solution is not necessary. Instead, Justesen imagines our project functioning as a treatment aimed at patients carrying OXA-48 after hospitalization. This would eliminate their carrier state, preventing unnecessary hospitalization in isolation, which is otherwise required whenever a carrier of OXA-48 producing bacteria is hospitalized, as well as transmission to other patients and people. Thereby, the reservoir of OXA-48 can be eliminated and treatment with common antibiotics can become useful once more. "),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null, "While Associate Professor Justesen provided invaluable knowledge on the day-to-day problems for clinicians, we also acknowledged that the input of relevant actors in the pharmaceutical industry was important to obtain a well-rounded project. Thus, we contacted SNIPR Biome, a young Danish biotech company. They utilize the CRISPR/Cas-technology to selectively kill pathogenic bacteria present in the human microbiome - a main goal much similar to ours. We visited them in Copenhagen and met with Head of Synthetic Biology and Bioinformatics, Eric van der Helm, and Research Scientist, Andreas Porse, to discuss the aspects of our conjugative CRISPR/Cas-system."),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement("p", null,
"After presenting our work to SNIPR Biome, we discussed some of the aspects of our project with them. We talked about the promising aspects of conjugation as a mode of delivery of our CRISPR/Cas-system. Until our meeting with SNIPR Biome, we had worked with dCas9, the catalytically inactive Cas9, as dCas9 could be a less cumbersome solution than Cas9 ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://dx.doi.org/10.1128%2FmBio.00928-13", target: "_blank" }, "Gomaa, A. A. et al. (2014) Programmable Removal of Bacterial Strains by Use of Genome-Targeting CRISPR-Cas Systems. mBio, 5(1), e00928-13. "))),
", which changes the microbiome. Even though we are working with plasmid-based DNA, the future potential of our project increases if implementable on chromosomal DNA. However, during our conversation they made a significant point convincing us that we should work on an alternative solution of assembling a system containing the catalytically active Cas9. Since Cas9 will knock down (and not only repress) the target gene, the elimination of resistant bacteria might be more efficient using Cas9 instead of dCas9. Furthermore, implementing Cas9 instead of dCas9 could be beneficial, as the system in this case does not need to be expressed simultaneously in order to inhibit the expression of the antibiotic resistance gene. However, Cas9 could result in other problems such as the accidental destruction of beneficial microbes ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://dx.doi.org/10.1128%2FmBio.00928-13", target: "_blank" }, "Gomaa, A. A. et al. (2014) Programmable Removal of Bacterial Strains by Use of Genome-Targeting CRISPR-Cas Systems. mBio, 5(1), e00928-13. "))),
". After eradication of the resistance genes by the action of a CRISPR/Cas9-system, treatment with the system can be terminated."),
React.createElement("p", null,
"Another interesting point brought to light by SNIPR Biome was the potential of the sequence targeted by our sgRNA mutating. Mutations in the target DNA will most likely prevent the recognition of the DNA by the sgRNA, in turn preventing cleavage/knockdown of the target gene. Regarding this issue, SNIPR Biome suggested implementing more sgRNAs, or a CRISPR/Cas array, into our system. Thus, we can potentially target different sequences in the target gene and lower the risk of the target gene not being recognized, despite mutations. This is especially relevant for",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Future_Teams", target: "_blank" }, " future work"),
". "),
React.createElement("p", { id: "no-bottom" }, "Our discussions with Associate Professor Justesen and SNIPR Biome provided us with indispensable knowledge about the implications of antibiotic resistance in the lives of Doctors, as well as relevant considerations regarding use of the CRISPR/Cas-technology. With these leading experts in the field, Conjugaid has been shaped - now it is our time to shape others. Click \u2018Next\u2019 to learn how we have engaged with the public beyond iGEM.")))),
React.createElement(WavesWidget_1.Waves, Object.assign({}, { waves: bottomWaves, top: "", concurrentZDelta: -1 })),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.IHP = IHP; //# sourceMappingURL=IHP.js.map } // default/src/components/Content/Future_Teams/Future_Teams.jsx $fsx.f[192] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const AccordionWidget_1 = $fsx.r(129); const rodal_1 = $fsx.r(132); const Buttons_1 = $fsx.r(104); const BulgeWidget_1 = $fsx.r(190); class FutureTeams extends React.Component {
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(191);
});
}
constructor(props) {
super(props);
this.state = { visibleEdu: false, visiblePizza: false };
window.document.title = `${props.title} | SDU-Denmark`;
}
show(tag) {
document.body.classList.add("rodal-open");
if (tag == "edu") {
this.setState({ visibleEdu: true });
}
else if (tag == "pizza") {
this.setState({ visiblePizza: true });
}
}
hide(tag) {
document.body.classList.remove("rodal-open");
if (tag == "edu") {
this.setState({ visibleEdu: false });
}
else if (tag == "pizza") {
this.setState({ visiblePizza: false });
}
}
render() {
let accordionDataProject = {
backgroundColor: "bg-blue-800",
title: "Possible Alterations and Additions to our Project Design",
itemTitleColor: "text-blue-800",
items: [
{
title: "OXA-48 producing Enterobacteriaceae and a sgRNA array",
content: `
One could test our CRISPR/(d)Cas9-system in actual OXA-48 carbapenemase producing Enterobacteriaceae. Downregulating resistance genes in these bacteria can improve and further support our proof-of-concept. For this purpose, we have made alignments of different OXA-48-like carbapenems. Consensus sequences from the OXA-48-like carbapenem resistance genes were also identified, enabling us to create a hypothetical sgRNA which could be incorporated into our plasmid containing the CRISPR/(d)Cas9-system.
The final sgRNA sequence made to target a consistent sequence common for a lot of OXA-48-like carbapenem resistance genes have been aligned. It was designed using the web tool CHOPCHOP <a href="https://academic.oup.com/nar/article/47/W1/W171/5491735/" target="_blank">Labun, K. et al. (2019). CHOPCHOP v3: expanding the CRISPR web toolbox beyond genome editing. Nucleid Acids Research, 47(W1), W171-W174.</a> .
5’-GAATATGGATATGCCCACAT-3’
We found several consistent areas amongst the OXA-48 gene sequences. This means it is possible to incorporate a sgRNA array into our system in the future and target different areas of the gene instead of one. Creating a sgRNA array lowers the risk of OXA-48 not being recognized, even if mutations in the gene occur. Thus, the final system would be more efficient in disrupting OXA-48 resistance genes. Furthermore, it could be investigated whether the final sgRNA sequence is present in the core genome of common Enterobacteriaceae in the human intestine in order to minimize interaction with off-target sequences <a href="https://www.ncbi.nlm.nih.gov/pubmed/10890397" target="_blank">Zhang, Z. et al. (2000). A greedy algorithm for aligning DNA sequences. J Comput Biol, 7(1-2), 203-14.</a> .
},
{
title: "Integration of our system in a conjugative plasmid",
content: `
Being able to assemble our designed CRISPR/Cas-system in a conjugative plasmid could also add other advantages to our project. This statement is based on our <a href="https://2019.igem.org/Team:SDU-Denmark/Model" target="_blanK">modeling results</a>. Integrating our designed CRISPR/Cas-system into the conjugative plasmid will increase the number of transconjugants leading to a higher success-rate regarding the propagation of the plasmid in a bacterial community.
},
{
title: "Work with flow cells",
content: `
Insight into the required dosage of bacteria given to patients carrying OXA-48-like carbepenemases producing Enterobacteriaceae can roughly be estimated by mimicking the gastrointestinal environment using flow cells <a href="https://www.ncbi.nlm.nih.gov/pubmed/29450193" target="_blank">Pedersen, R. M. et al. (2018). A Method for Quantification of Epithelium Colonization Capacity by Pathogenic Bacteria. Front Cell Infect Microbiol, 8:16. </a> . Intestinal epithelial cells have to be grown to a confluent monolayer followed by the addition of a flow of an intestine-like medium in the chamber. OXA-48-like carbepenemase-producing Eenterobacteriaceae can then be seeded onto the artificial intestine followed by inoculation of donor bacteria. Comparing the remaining bacteria to a control experiment will then give a rough estimate on how many OXA-48-like carbepenemases producing Enterobacteriaceae have been affected by the CRISPR/Cas-system. Such an experiment could add another opportunity to study the conjugation efficiency of a drug candidate like ours in a more in vivo-like environment.
},
{
title: "Using E. coli Nissle as a chassis",
content: `
When ingesting bacteria it is imperative that the intestine is not colonized, since this could cause diseases. To prevent this from happening, non-pathogenic and probiotic bacteria such as E. coli Nissle could be used as donor bacteria in the CRISPR/Cas-system <a href="https://www.ncbi.nlm.nih.gov/pubmed/25380207" target="_blank">Różańska, D. et al. (2014). The role of Escherichia coli strain Nissle 1917 in the gastro-intestinal diseases. Postepy Hig Med Dosw, 68, 1251-1256.</a> .
},
{
title: "Implementation of a kill-switch",
content: `
A heat sensitive kill-switch like the one used by the <a target="_blank" href="https://2017.igem.org/Team:Michigan">iGEM team from Michigan made in 2017</a> (BBa_K2301000 - BBa_K2301003) could be implemented in our system. As such, any possible environmental risks could be reduced, since the donor bacteria would be killed at temperatures below 37 ℃.
},
]
};
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "To Future iGEM Teams References",
content: `
- <a href="https://academic.oup.com/nar/article/47/W1/W171/5491735" target="_blank" class="references">Labun, K. et al. (2019). CHOPCHOP v3: expanding the CRISPR web toolbox beyond genome editing. Nucleid Acids Research, 47(W1), W171-W174.</a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/10890397" target="_blank" class="references">Zhang, Z. et al. (2000). A greedy algorithm for aligning DNA sequences. J Comput Biol, 7(1-2), 203-14.</a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/29450193" target="_blank" class="references">Pedersen, R. M. et al. (2018). A Method for Quantification of Epithelium Colonization Capacity by Pathogenic Bacteria. Front Cell Infect Microbiol, 8:16. </a>
- <a href="https://www.ncbi.nlm.nih.gov/pubmed/25380207" target="_blank" class="references">Różańska, D. et al. (2014). The role of Escherichia coli strain Nissle 1917 in the gastro-intestinal diseases. Postepy Hig Med Dosw, 68, 1251-1256. </a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "futureTeams" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "To Future iGEM Teams", img: "
", fromColor: "#1D3C4D", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\"Even if I knew that tomorrow the world would go to pieces, I would still plant my apple tree.\""),
React.createElement("p", { className: "quotemaster" }, "- MARTIN LUTHER, religious reformer"),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null, "Hello future iGEM team! Are you ready to begin your iGEM journey? You are going to have an absolutely amazing time and you will learn so much."),
React.createElement("p", null, "The world is facing many problems in desperate need of solutions. Our team set out to help solve just one of them. With Conjugaid, we have planted a seed in the hopes of it growing into a beautiful, fruit-bearing tree. But we cannot do this alone - we need the help of others to get there in time."),
React.createElement("p", null, "In this section you will find some ideas for further development of our project, Conjugaid, and tips for creating educational material. We will also be giving you a super-secret tip which totally saved our team when the days got a little rough."),
React.createElement("p", null, "One thing we are sure of when it comes to working in the laboratory is that you can always do more. If your team has been inspired by Conjugaid and you want to improve it, here are some suggestions as to how you could do that.")),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionDataProject })),
React.createElement("div", { className: "row" },
React.createElement("h3", null, "A recipe for a great educational material:"),
React.createElement("p", null,
"If your team wants to create educational material as a part of your iGEM project, look no further! We have created, tried and tested a ",
React.createElement("span", null,
React.createElement("button", { className: "text-blue-600", onClick: this.show.bind(this, "edu") }, "10-step recipe")),
" that should help you get started."),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement(rodal_1.default, { closeOnEsc: true, customStyles: {
width: "80vw",
height: "80vh"
}, visible: this.state.visibleEdu, onClose: this.hide.bind(this, "edu") },
React.createElement("div", { className: "rodal-box" },
React.createElement("div", { className: "border-solid border-gray-700 border-b " },
React.createElement("h3", null, "10 steps to creating great educational material")),
React.createElement("ol", null,
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Find out your target group"),
React.createElement("p", { className: "italic" }, "It is imperative when creating educational material to know who your students are. Teaching children in elementary school about microbes requires something completely different than teaching high schoolers does.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Talk to your target group (and their teachers)"),
React.createElement("p", { className: "italic" }, "Before you can develop great materials, you need to know the needs of the recipients. What do they lack, and what do they already have in abundance? Your way forward is to fill out the gaps.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Tell a story"),
React.createElement("p", { className: "italic" }, "One important rule of thumb about education is that learning is eased through storytelling. We taught antibiotic resistance through the story of Peter - how will you tell yours?")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Plan out everything in detail"),
React.createElement("p", { className: "italic" },
"Before you can start writing and designing your educational material, you need a solid plan. How many subjects do you wish to cover - and what should they be? What kind of illustrations do you need? Sort everything into chapters and write down what they should contain. You could be inspired by different teaching and learning models, like we did with the ",
React.createElement("a", null, "5E Teaching Model"),
".")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Write, write and rewrite..."),
React.createElement("p", { className: "italic" }, "Start writing your materials! And when you think you are done, look at it again - is it really done? Great! Now get someone else to read and comment on it, and then rewrite the whole thing to perfection.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Make it look stunning!"),
React.createElement("p", { className: "italic" }, "This is where you start taking your educational material to the next level. Does someone on your team love drawing? Ask them to create the illustrations and get designing - the options are endless, so have fun with it!")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Finishing touches"),
React.createElement("p", { className: "italic" }, "When your educational material is all done - written, illustrated and designed - forward it to a group of people and get their final comments on it. Make sure to implement these comments before testing your material.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, " Survey before..."),
React.createElement("p", { className: "italic" }, "Congratulations! You have now created your very own educational material - that is amazing. All that is left now is to test it out on your target group and find out how they like it. But before you can test your material and evaluate it, you need to survey your test group. That way, you can compare how your educational material has directly affected the people using it.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "Put it to the test"),
React.createElement("p", { className: "italic" }, "Go on - do it! You have nothing to worry about, you have worked so hard and put so much effort into it, we are sure your educational material will do amazingly with your test group.")),
React.createElement("li", null,
React.createElement("h5", { className: "font-bold" }, "\u2026 And evaluate after!"),
React.createElement("p", { className: "italic" }, "This is the last, but definitely not least important, step of this recipe. You have created this amazing educational resource, and you want more people to notice and use it, right? In order for them to do so, they need to see results. Ask your test group any questions that might be relevant for you to properly evaluate how your educational material did in practice."))))),
React.createElement("h3", null, "A recipe for something else that\u2019s also great\u2026"),
React.createElement("p", null,
"Perhaps you\u2019re a new iGEM team, innocent and unaware of the fact that you will soon be busier than ever. Perhaps your team has been going strong for some months now, you\u2019re starting to feel the pressure and now you\u2019ve come to us for advice. No matter the situation, you\u2019ve come to the right place! ",
React.createElement("span", null,
React.createElement("button", { className: "text-blue-600", onClick: this.show.bind(this, "pizza") }, "This is a recipe for the pizza that will save your lives when times are tough."))),
React.createElement("div", { className: "img_container" },
React.createElement("img", { className: "", src: "
" })),
React.createElement(rodal_1.default, { closeOnEsc: true, customStyles: {
width: "80vw",
height: "80vh"
}, visible: this.state.visiblePizza, onClose: this.hide.bind(this, "pizza") },
React.createElement("div", { className: "rodal-box" },
React.createElement("div", { className: "border-solid border-gray-700 border-b " },
React.createElement("h3", null, "6 steps to creating the greatest pizza ")),
React.createElement("p", null, "Pi(GEM)zza (12 people/3 pizzas):"),
React.createElement("p", null, "Prep-time: 30 minutes"),
React.createElement("p", null, "Cooking time: 15 minutes"),
React.createElement("p", null, "Measuring units: Metric (as scientists we like to use the proper system)"),
React.createElement("p", { className: "italic" }, "Ingredients"),
React.createElement("ul", null,
React.createElement("li", null, "3 x Store bought pizza dough (you think you have time to make your own while doing iGEM? Just you wait and see\u2026)"),
React.createElement("li", null, "3 x Garlic cream cheese spread"),
React.createElement("li", null, "450 g shredded cheddar cheese"),
React.createElement("li", null, "1.5 kg potatoes, washed and thinly sliced"),
React.createElement("li", null, "3 red onions, thinly sliced"),
React.createElement("li", null, "3 packets of pepperoni"),
React.createElement("li", null, "A few sprigs of fresh rosemary"),
React.createElement("li", null, "Sea salt"),
React.createElement("li", null, "A teaspoon team-spirit and iGEM excellence")),
React.createElement("p", { className: "italic" }, " Directions"),
React.createElement("ol", null,
React.createElement("li", null, "Roll out the pizza doughs and spread one container of garlic cream cheese on each pizza"),
React.createElement("li", null, "Sprinkle 150g shredded cheddar cheese on each pizza"),
React.createElement("li", null, "Distribute the potato slices on the pizzas, then the pepperoni and then the slices of red onion"),
React.createElement("li", null, "Add a few sprigs of fresh rosemary to each pizza and sprinkle with sea salt, team-spirit and iGEM excellence to taste"),
React.createElement("li", null, "Bake in a 200\u00B0C pre-heated oven for 12-15 minutes or until golden and crispy"),
React.createElement("li", null, "Serve to your eager and hungry iGEM-team and prepare for everyone to compliment you on your great cooking skills!")))))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.FutureTeams = FutureTeams; //# sourceMappingURL=Future_Teams.js.map } // default/src/components/widgets/BulgeWidget.jsx $fsx.f[190] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const _ = $fsx.r(44); const $ = $fsx.r(102); class BulgeWidget extends React.Component {
constructor(props) {
super(props);
this.topWaveRef = React.createRef();
this.listener = _.throttle(this.updateDimensions.bind(this), 50);
this.state = { overlayHeight: "100%", captionBottom: 0 };
this.imageRef = React.createRef();
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(189);
});
}
componentDidMount() {
window.addEventListener('scroll', this.listener);
window.addEventListener('resize', this.listener);
this.imageRef.current.onload = this.updateDimensions.bind(this);
this.imageRef.current.src = this.props.img;
}
componentWillUnmount() {
window.removeEventListener('scroll', this.listener);
window.removeEventListener('resize', this.listener);
}
inViewport(ref) {
var H = $(window).height(), r = ref.getBoundingClientRect(), t = Math.min(0, r.top), b = r.bottom - 155;
return Math.max(0, t > 0 ? H - t : (b < H ? b : H));
}
updateDimensions() {
if (!this.imageRef.current)
return;
let height = this.imageRef.current.getBoundingClientRect().height;
if (height == 0) {
setTimeout(this.updateDimensions.bind(this), 20);
return;
}
let ref = this.topWaveRef.current;
if (ref) {
let window_offset = this.inViewport(ref);
let deg = window_offset * 0.0;
let fromColor = this.props.fromColor ? this.props.fromColor : "red";
let toColor = this.props.toColor ? this.props.toColor : "#ffda00";
this.setState({
overlayHeight: window_offset,
captionBottom: window_offset / 4,
gradient: `linear-gradient(${deg}deg, ${fromColor}, ${toColor})`
});
}
}
render() {
return (React.createElement("div", { className: "top-wave noselect", ref: this.topWaveRef, style: {
backgroundImage: this.state.gradient,
} },
React.createElement("img", { alt: "", ref: this.imageRef }),
React.createElement("figcaption", { className: "caption", style: {
bottom: this.state.captionBottom
} },
React.createElement("h1", { className: "noselect" }, this.props.title)),
React.createElement("span", { className: "overlay", style: {
height: this.state.overlayHeight,
} },
React.createElement("svg", { version: "1.1", id: "circle", xmlns: "http://www.w3.org/2000/svg", x: "0px", y: "0px", viewBox: "0 0 500 250", enableBackground: "new 0 0 500 250", preserveAspectRatio: "none" },
React.createElement("path", { fill: "#FFFFFF", d: "M250,246.5c-97.85,0-186.344-40.044-250-104.633V250h500V141.867C436.344,206.456,347.85,246.5,250,246.5z" })))));
}
} exports.BulgeWidget = BulgeWidget;
//# sourceMappingURL=BulgeWidget.js.map } // default/src/components/Content/Achievements/Achievements.jsx $fsx.f[194] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); class Achievements extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(193);
});
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "achievements" },
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Achievements"),
React.createElement("div", { className: "under-rubrik" }, "What have we accomplished?"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "Throughout our project - Conjugaid - our team has achieved many things. This section serves as an overview showing you exactly which medal-criteria we have fulfilled."))),
React.createElement("div", { className: "row" },
React.createElement("div", { className: "achievement-border bronze-border" },
React.createElement("img", { className: "medal-img", src: "
" }),
React.createElement("p", { className: "medal-text bronze-text" }, "Bronze Criteria")),
React.createElement("ol", null,
React.createElement("li", null,
"Registration and Giant Jamboree Attendance ",
React.createElement("span", { className: "flueben" }, "\u2713")),
React.createElement("li", null,
"Competition Deliverables: Convince the judges that you have completed the following Competition Deliverables: ",
React.createElement("br", null),
React.createElement("p", { className: "ml-12" },
"#1 Wiki ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" "),
React.createElement("p", { className: "ml-12" },
"#2 Poster ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" "),
React.createElement("p", { className: "ml-12" },
"#3 Presentation ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" "),
React.createElement("p", { className: "ml-12" },
"#4 Judging Form ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ")),
React.createElement("li", null,
"Attributions: Convince the judges that you have completed Competition Deliverable #5 ",
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Attributions" }, "Attributions "),
". ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" "),
React.createElement("li", null,
"Project Inspiration and Description: Convince the judges that you have completed Competition Deliverable #8 ",
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Description" }, " Project Inspiration and Description"),
". ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" "),
React.createElement("li", null,
"Characterization: Convince the judges that you have added quantitative experimental characterization data to an existing Part from the Registry of Standard Biological Parts. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic" },
"Our team has characterized the parts ",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_J01003", target: "_blank" }, "BBa_J01003"),
" and ",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_I714030", target: "_blank" }, "BBa_I714030"),
" by measuring transfer efficiency using different conjugative plasmids.")))),
React.createElement("div", { className: "row" },
React.createElement("div", { className: "achievement-border silver-border" },
React.createElement("img", { className: "medal-img", src: "
" }),
React.createElement("p", { className: "medal-text silver-text" }, "Silver Criteria")),
React.createElement("ol", null,
React.createElement("li", null,
React.createElement("a", { target: "_blank", href: "" }, "Validated Part"),
": Convince the judges that at least one new BioBrick Part of your own design that is related to your project works as expected. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic text-red-600" },
"Our team has created the part ",
React.createElement("a", { href: "http://parts.igem.org/Part:BBa_K3044027", target: "_blank" }, "BBa_K3044027"),
", which is a sgRNA-Cas9 system targeting GFP.")),
React.createElement("li", null,
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Collaborations" }, "Collaborations"),
": Convince the judges you have significantly worked with one (or more) currently registered 2019 iGEM team(s) in a meaningful way. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic" },
"Throughout our project, our team has worked with the following iGEM teams: ",
React.createElement("a", { href: "https://2019.igem.org/Team:DTU-Denmark", target: "_blank" }, "DTU Biobuilders"),
", ",
React.createElement("a", { href: "https://2019.igem.org/Team:UCopenhagen", target: "_blank" }, "UCopenhagen"),
", ",
React.createElement("a", { href: "https://2019.igem.org/Team:Amazonas-Brazil", target: "_blank" }, " Amazonas-Brazil"),
", ",
React.createElement("a", { href: "https://2019.igem.org/Team:Lund", target: "_blank" }, "Lund"),
", ",
React.createElement("a", { href: "https://2019.igem.org/Team:TAS_Taipei", target: "_blank" }, "TAS_Taipei"),
", ",
React.createElement("a", { href: "https://2019.igem.org/Team:Costa_Rica", target: "_blank" }, "Costa Rica"),
" & ",
React.createElement("a", { href: "https://2019.igem.org/Team:Tuebingen", target: "_blank" }, "Tuebingen"),
".")),
React.createElement("li", null,
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices" }, "Human Practices"),
": Convince the judges you have thought carefully and creatively about whether your work is responsible and good for the world. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic" }, "Our team has created and tested a lesson-plan for high school students. Furthermore, we have engaged the public as well as legislators in regards to synthetic biology and antibiotic resistance.")))),
React.createElement("div", { className: "row" },
React.createElement("div", { className: "achievement-border gold-border" },
React.createElement("img", { className: "medal-img", src: "
" }),
React.createElement("p", { className: "medal-text gold-text" }, "Gold Criteria")),
React.createElement("ol", null,
React.createElement("li", null,
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices" }, "Integrated Human Practices"),
": Expand on your silver medal activity by demonstrating how you have integrated the investigated issues into the purpose, design, and/or execution of your project. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic" }, "We have continuously altered our project purpose, design and execution after discussing our ideas with Professor in clinical microbiology at our local hospital Ulrik S. Justesen and with SNIPR Biome.")),
React.createElement("li", null,
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Model" }, " Model Your Project"),
": Convince the judges that your project's design and/or implementation is based on insight you have gained from modeling. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic" }, "Through modelling, we have gained insight about the optimal design and implementation of our conjugation system. Efficiency of delivery of our CRISPR/(d)Cas9-system is highest if placed on a single conjugative plasmid. Furthermore, the number of recipient bacteria will increase this way."))),
React.createElement("ol", { start: 4 },
React.createElement("li", null,
React.createElement("a", { target: "_blank", href: "https://2019.igem.org/Team:SDU-Denmark/Demonstrate" }, "Demonstration of Your Work "),
": Convince the judges that your engineered system works. ",
React.createElement("span", { className: "flueben" }, "\u2713"),
" ",
React.createElement("p", { className: "italic" }, "Based on our results within our two systems, we determined conjugative delivery and CRISPR/Cas-activity to be successful, thus demonstrating our work."))))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Achievements = Achievements; //# sourceMappingURL=Achievements.js.map } // default/src/components/Content/People_Involved/Collaborations/Collaborations.jsx $fsx.f[195] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const AccordionWidget_1 = $fsx.r(129); const Buttons_1 = $fsx.r(104); const nuka_carousel_1 = $fsx.r(168); class Collaborations extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
this.carousel = React.createRef();
this.state = {
carouselHeight: 0
};
}
componentDidMount() {
let carousel = this.carousel.current;
if (carousel) {
let constant = 0.5625;
let parent = carousel.frame.parentElement;
let parentRect = parent.getBoundingClientRect();
let newheight = parentRect.width * constant;
this.setState({
carouselHeight: newheight
});
}
}
render() {
let accordionData = {
backgroundColor: "bg-green-400",
title: "Collaboration",
itemTitleColor: "text-green-800",
items: [
{
title: "LGBTQ+ Visbillity",
content: `
During the Danish iGEM Weekend the idea came up for our team to participate in the Copenhagen Pride Parade with Ovulaid, the iGEM team from University of Copenhagen. On a lovely afternoon in mid-August, our team walked with Ovulaid in the Pride Parade as part of the University of Copenhagen’s float. We did this to promote more diversity in science.
<img id="box-img" src="
"> </img>
`
},
{
title: "Attending Meetups",
content: `
DTU BioBuilders, BioBrick Tutorial, April 2019
From the 12th to the 14th of April 2019, we traveled to the Technical University of Denmark, where DTU BioBuilders are based, in order to participate in their 7th annual BioBrick Tutorial. Here, we learned the basics of how to work with iGEM BioBricks. This was also the first time we were introduced to other iGEM teams, and it was a pleasure to get to know the teams of DTU BioBuilders, Copenhagen, Stockholm, Chalmers, Uppsala, Oslo, Lund and Helsinki.
<img class="f-w-img" src="
"> </img>
iGEM Team Copenhagen, Nordic iGEM Conference, May 2019
From the 17th to the 19th of May 2019, we visited the University of Copenhagen and their iGEM team, Ovulaid. Here, we participated in this years’ Nordic iGEM Conference, an annual event consisting of workshops on multiple subjects (this year science communication and sustainability was on the programme, amongst other things). Besides having a great time with the other nordic iGEM teams, we left the conference with the award for “Best Poster”.
<img class="f-w-img" src="
"> </img>
`
},
{
title: "iGEM Team Amazonas Brazil",
content: `
In July 2019 we were contacted by the iGEM team of Amazonas (Brazil). They reached out to iGEM teams all over the world through a common chat (‘Sharing iGEM Ideas’). Here, they asked if anyone had assembled a plasmid containing dCas9, since they required it for their project. At that time we were hard at work trying to design and assemble it ourselves, so we reached out to them and let them know that we would contact them when we succeeded. A few weeks later we had designed, assembled and sequenced our plasmid containing dCas9, and could send it their way.
<img id="box-img" src=""></img>
`
},
{
title: "DTU BioBuilders",
content: `
After discussing our individual teams’ processes at the Danish iGEM Weekend, the DTU BioBuilders contacted us for technical assistance. Since they ran out of pSB1K3 backbone, we sent some of our pSB1K3 part from the iGEM kit to them so they could continue their work.
`
},
{
title: "iGEM Team Lund",
content: `
When beginning our work within modelling, we stumbled upon some initial troubles. We remembered talking to a Arvid Larsson, modelling-extraordinaire from the 2019 iGEM team Lund, at both the BioBrick Tutorial and the Nordic iGEM Conference. After reaching out to him we had a discussion via Skype in which we talked all things modelling. Arvid provided us with valuable advice and support in designing our modeling system.
<img id="box-img" src="
"></img>
`
},
{
title: "iGEM Teams’ Tas_Taipei, Costa Rica & Tuebingen iGEMxSDGs Instagram Challenge",
content: `
For this years’ iGEM Competition, the teams of TAS_Taipei (Taiwan), Costa Rica and Tuebingen (Germany) collaborated to create the iGEMxSDGs Instagram Challenge. This challenge took advantage of the fact that each iGEM team communicates continuously with their communities, and used this opportunity to promote the UN’s 17 SDGs. The SDGs can only be achieved if we work on them collaboratively on a global scale, and what better way to help facilitate this than by working on the SDGs within iGEM.
To participate in the challenge, each team had to select up to four SDGs targeted by their project and post them on Instagram with a short description. The challenge was then spread throughout the iGEM community when each team challenges four other teams to participate.
Our team naturally participated and posted the following four SDGs, which we directly target in our project:
- SDG No. 3: Good health and well-being
- SDG No. 4: Quality education
- SDG No. 12: Responsible consumption and production
- SDG No. 17: Partnerships for the goals
You can read more about our work with the SDGs <a> here </a>.
`
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "collaborations" },
React.createElement("div", { className: "content pb-20" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Collaborations"),
React.createElement("div", { className: "under-rubrik" }, "Teamwork makes the dream work"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, "Our team has worked with iGEM teams from all over the world throughout this year\u2019s competition - and it has been so much fun! Keep reading to learn what exactly we\u2019ve been up to..."))),
React.createElement("div", { className: "row" },
React.createElement("h2", { className: "whiter-border" }, "The Danish iGEM Weekend"),
React.createElement("p", null, "In recent years, Danish iGEM teams have participated in both the BioBrick Tutorial at the Technical University of Denmark and the Nordic iGEM Conference (NiC). Both of these meetups are international and participating teams come from all over Scandinavia. After meeting the two other Danish teams (Ovulaid from University of Copenhagen and DTU Biobuilders from DTU) we wished to further strengthen the bond between the Danish iGEM teams. This lead to us organizing an iGEM meetup for Danish teams; the Danish iGEM Weekend. The meetup tackled subjects such as wiki-building and public presentation skills. Through workshops and talks hosted by both our own team members and external mediators, all teams received the proper foundation needed to start building and telling the story of their iGEM projects. Ovulaid and the DTU Biobuilders returned home after our meetup and felt prepared to begin creating their wiki- and Jamboree presentation-storylines. Furthermore, we hope to have started a new tradition of hosting the Danish iGEM Weekend for future iGEM teams from SDU. The bond between the Danish iGEM teams has been strengthened, and our aim is to carry this strong bond into the future. "),
React.createElement(nuka_carousel_1.default, { ref: this.carousel, className: "mx-auto my-10", width: "100%", dragging: true, style: {
height: this.state.carouselHeight,
} },
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" })),
React.createElement("p", null, "Read more about our other collaborations in the foldouts below:")),
React.createElement(AccordionWidget_1.AccordionWidget, Object.assign({}, { model: accordionData }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Collaborations = Collaborations; //# sourceMappingURL=Collaborations.js.map } // default/src/components/Content/People_Involved/TeamPage/TeamPage.jsx $fsx.f[199] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const react_lazy_load_image_component_1 = $fsx.r(196); $fsx.r(197); class TeamPage extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(198);
});
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "teamPage" },
React.createElement("div", { className: "content pb-20" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "rubrik" }, "Team"),
React.createElement("div", { className: "under-rubrik" }, "Individual presentations"),
React.createElement("div", { className: "manchet" },
React.createElement("p", { className: "" }, " Welcome to the Team & Supervisors page! Our team consists of 11 students from two different faculties and four different studies, and we are supported by three amazing supervisors. Since beginning our iGEM journey in December 2018, we have developed an unbreakable iGEM bond and gotten to know each other - we think you deserve to get to know us on a more personal level, too. "))),
React.createElement("div", { className: "row" },
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Anne Sofie L\u00F8gstrup "),
React.createElement("p", null, "Study: Biomedicine"),
React.createElement("p", null, "E-mail: anhen17@student.sdu.dk"),
React.createElement("p", null, "Hi folks! I\u2019m Anne Sofie, and for the past 10 months I\u2019ve been working as a middleman between our donor- and acceptor-bacteria, forcing them to be friendly and talk to each other via conjugation. When not in the lab being a bacterial couples therapist, my biggest challenge has been learning sarcasm - I mean, what kind of mysterious language is that?! Luckily my teammates learned to use jazz hands when being sarcastic, which makes it WAY easier to translate what they\u2019re saying! "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Annemette Pr\u00E6stegaard Thulesen "),
React.createElement("p", null, "Study: Biochemistry and molecular biology"),
React.createElement("p", null, "E-mail: anthu17@student.sdu.dk"),
React.createElement("p", null, "Hello! I\u2019m Annemette - a glasses-wearing, volleyball-playing, impro-acting, bacteria-loving, sock-knitting ginger. Throughout the entirety of our iGEM project I\u2019ve practically been camped out in our laboratory, doing bacteriamazing work day in and day out! "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Catharina Bang Jensen"),
React.createElement("p", null, "Study: Biochemistry and molecular biology"),
React.createElement("p", null, "E-mail: catje17@student.sdu.dk"),
React.createElement("p", null, "Hejsa! I\u2019m Catharina, and I enjoy spending time in the lab - even when it doesn\u2019t work! This project has been hard to complete without a little bit of cash, for which reason I have been fundraising. I\u2019ve also been optimizing our lab-results in the computational group, but you will primarily find me in the laboratory where my inner Lab Queen will be showing her magic by using her mad pipetting skills and wearing an oversized lab coat all summer long."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Christian Gejl Christensen"),
React.createElement("p", null, "Study: Biomedicine"),
React.createElement("p", null, "E-mail: chrgejl@hotmail.com"),
React.createElement("p", null, "Heyo! I\u2019m Christian, but don\u2019t confuse my name with having the virtues of one. My parents have always told my that nothing is impossible, but I did nothing besides studying biomedicine all day until I joined this wonderful iGEM team. Those slacking days are over as I am now wikipediating in our wiki group, getting \u2018em $$$\u2019s in our fundraising group and doing human stuff in our human practices group."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Ditlev Hartmann Bornebusch "),
React.createElement("p", null, "Study: Life Science Engineering and Informatics (OMICS)"),
React.createElement("p", null, "E-mail: deltaetabeta@gmail.com"),
React.createElement("p", null, "Hi! I\u2019m Ditlev, the engineer of the team! I am a professional computer wizard and Life Science apprentice, who aspires to harness life sorcery with digital magic. I thrive the most in a programming environment with unlimited coffee, funky disco music, multiple monitors, a high-speed internet connection and at minimum a machine with 16GB RAM. Oh, and I also built most of the wiki from my new home in China!"))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Emil Birch Christensen"),
React.createElement("p", null, "Study: Biochemistry and molecular biology"),
React.createElement("p", null, "E-mail: emilc16@student.sdu.dk"),
React.createElement("p", null, "Hello, Emil here. I recently finished my BA in Biomedicine and have now moved on to a Masters in biochemistry and molecular biology. Since I have already used up all of my young and fruitful years studying, I thought I might as well add fuel to the flame and join the iGEM team. You\u2019ll find me setting stuff on fire in the lab, doing wizard stuff in the wiki-group and asking people for their money in the fundraising group."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Emma Emilie Andersen"),
React.createElement("p", null, "Study: Biology"),
React.createElement("p", null, "E-mail: emman17@student.sdu.dk"),
React.createElement("p", null, "Hi there! I\u2019m Emma, the biologist of the team. Obsessed with loads of things that nobody else on the team seem to care about (even though they totally should!), I make sure everyone notices all of the wiggly worms and precious plants we pass on our way. Throughout our project I\u2019ve been working with humans in Human Practices, making agar plates and overnight cultures as a servant to our \u201Creal\u201D lab-team, cashing checks in fundraising and getting followers on social media (#ShoutoutToiGEM). "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Henrik Frederik Bekkevold Johansen"),
React.createElement("p", null, "Study: Biochemistry and molecular biology"),
React.createElement("p", null, "E-mail: hejoh15@student.sdu.dk"),
React.createElement("p", null, "Hey y\u2019all! I\u2019m Henrik, a former medical student who turned to the dark side. I now spend my time with biochemistry and molecular biology and I gave my soul to the Dark Lord in exchange for being a part of iGEM. I'm the Protector of the human practices mailbox, Writer of e-books, Queen of shade and sarcasm, bound by oath to the lab team, Purifier of plasmids, Khaleesi of the Great Agar Sea and Mother of plates. "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Jacob Fredegaard Hansen"),
React.createElement("p", null, "Study: Biochemistry and molecular biology"),
React.createElement("p", null, "E-mail: jaha217@student.sdu.dk"),
React.createElement("p", null, "Hi there! My name is Jacob, and in our iGEM project I\u2019m hustling in human practices, with fundraising and with social media (noticed all of the amazing, high-quality photos on our SoMe-pages - and these portraits of our beautiful team? Yup, I took those!). However, my biggest talent is making everyone laugh with my dad-jokes! Hit me up if you wanna hear one, I got plenty."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Stine Degn Espersen"),
React.createElement("p", null, "Study: Biochemistry and molecular biology"),
React.createElement("p", null, "E-mail: stesp17@student.sdu.dk"),
React.createElement("p", null, "Hi, what\u2019s up? My name is Stine, and I\u2019ve always been fascinated by how synthetic biology can be used to our advantage *insert evil german scientist laugh*. In this regard, I primarily work in the lab, but I\u2019m also working on making our project look good by designing beautiful wiki-, presentation- and poster-illustrations. "))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { src: "
", effect: "blur" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("h5", { className: "font-black text-lg" }, "Thea Amalie Hvidtfeldt"),
React.createElement("p", null, "Study: Biomedicine"),
React.createElement("p", null, "E-mail: thvid17@student.sdu.dk"),
React.createElement("p", null, "Hi everyone! I\u2019m Thea and during iGEM I\u2019ve been working with a lot of (E.) Cool(i) bacteria in the lab. Since I am resistant to the fact that I needed a lot of food for long iGEM days, I\u2019ve been very sensitive to canteen-trip invitations. However, when nutrients (primarily sugar and coffee) are available, my energy rises at an exponential rate!"))),
React.createElement("h2", { className: "whiter-border" }, "Supervisors"),
React.createElement("p", null, "A special thanks to our supervisors: "),
React.createElement("h5", null,
React.createElement("span", null, "Mikkel Girke J\u00F8rgensen"),
", associate Professor at SDU\u2019s Department of Biochemistry and Molecular Biology"),
React.createElement("p", null, "Mikkel has been the primary supervisor throughout our project, and he has been responsible for handling iGEM on SDU for many years - so he\u2019s really good at it! Mikkel has been of assistance to us within every aspect of our project, but always makes it a point to never tell us no. Whatever we wanted to do, we could do - Mikkel has merely been gently guiding us."),
React.createElement("h5", null,
React.createElement("span", null, "Felix Boel Pedersen"),
", MSc student"),
React.createElement("p", null, "Like Frederik, Felix is a former iGEM participant from team SDU 2017. Felix has been of assistance in all areas of our project, from the beginning till the end, but his wisdom when it comes to creating the perfect iGEM wiki has been especially influential."),
React.createElement("h5", null,
React.createElement("span", null, "Frederik Bartholdy Flensmark Neergaard"),
", MSc student"),
React.createElement("p", null, "Frederik is a former iGEM participant from team SDU 2017. From teaching us general cloning in the very beginning to helping perfect our wiki-text just before the freeze, Frederik has been of great help to our team throughout our entire project."),
React.createElement(react_lazy_load_image_component_1.LazyLoadImage, { className: "mt-10 rounded-lg shadow-xl", src: "
", effect: "blur" }))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.TeamPage = TeamPage; //# sourceMappingURL=TeamPage.js.map } // default/src/components/Content/Project/ProjectOverview.jsx $fsx.f[201] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const BulgeWidget_1 = $fsx.r(190); const isMobile_1 = $fsx.r(58); const eq1 = `\\\frac{dTN}{dt}=\\\gamma R\\\alpha_N(CN+TCN)-\\\gamma TN[TC+(\\\alpha_C+\\\alpha_C_N)(CN+TCN)]`; const m_break = isMobile_1.default ? "\\\\" : ""; class ProjectOverview extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(200);
});
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "Project References",
content: `
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4888801/" target="_blank" class="references">Munita, J. M. and Arias, C. A. (2016). Mechanisms of Antibiotic Resistance. Microbiol. Spectr., 4(2), April. </a>
- <a href="https://www.researchgate.net/publication/298640728_Laboratory_Detection_and_Clinical_Implication_of_Oxacillinase-48_like_Carbapenemase_The_Hidden_Threat" target="_blank" class="references">Yamuna, D. et al. (2016). Laboratory Detection and Clinical Implication of Oxacillinase-48 like Carbapenemase: The Hidden Threat. J. of global infectious diseases, 8, 41-50. </a>
- <a href="https://www.msdmanuals.com/professional/infectious-diseases/gram-negative-bacilli/escherichia-coli-infections" target="_blank" class="references">Bush, L. M. and Perez, M. T. (2018). Escherichia coli Infections. MSD Manual. Accessed October 1 2019.</a>
- <a href="https://www.sciencedirect.com/science/article/pii/S0140673603136446?fbclid=IwAR3UwlyPW-_xdzJDY39shKuxE5ZkApXBIQUuZGz7A0--8vf7LhxjO2viFF4" target="_blank" class="references">Vincent, J. L. (2003). Nosocomial infections in adult intensive-care units. The Lancet, 361(9374), 2068-2077.</a>
- <a href="https://www.sciencedirect.com/science/article/pii/S1438422113000167?via%3Dihub&fbclid=IwAR2I23U_mj_zfHGtWmjZdpso8TvCpv2_KWxWvZayRSuHsNd4HoaD0Zxe2Cs" target="_blank" class="references">Carattoli, A. (2013). Plasmids and the spread of resistance. Int. J. of Med. Microbiology, 303(6-7), 298-304.</a>
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4378521" target="_blank" class="references">Ventola, C. L. (2015). The Antibiotic Resistance Crisis, Part 1: Causes and Threats. PT, 40(4), 277-283.</a>
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5330996" target="_blank" class="references">Tran, K. et al. (2017). The Effect of Hospital Isolation Precautions on Patient Outcomes and Cost of Care: A Multi-Site, Retrospective, Propensity Score-Matched Cohort Study. J Gen Intern Med., 32(3), 262-268. </a>
- <a href="https://www.sciencedirect.com/science/article/pii/S1021949819300122" target="_blank" class="references">Ding, R-x. et al. (2019). Revisit gut microbiota and its impact on human health and disease. J of Food and Drug Analysis, 3, 623-631. </a>
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4343198" target="_blank" class="references">Hsu, P. D. et al. (2014). Development and Applications of CRISPR-Cas9 for Genome Engineering. Cell, 157(6), 1262-1278.</a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "project-overview" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "Project", img: "
", fromColor: "#1D3C4D", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CReligion is a culture of faith; science is a culture of doubt.\u201D"),
React.createElement("p", { className: "quotemaster" }, "\u2015 Richard P. Feynman, American physicist"),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null,
"Antibiotic resistance is a global problem, and different bacterial strains and resistances influence different regions of the world ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.who.int/glass/resources/publications/early-implementation-report-2017-2018/en/", target: "_blank" }, "Global antimicrobial resistance surveillance system (GLASS) report: early implementation 2017-2018. Geneva: World Health Organization; 2018. Licence: CC BY-NC-SA 3.0 IGO."))),
". In our hometown, Odense, oxacillinase-48 (OXA-48) producing Enterobacteriaceae are of special relevance, according to Associate Professor Ulrik S. Justesen, Clinical Microbiologist at University Hospital Odense."),
React.createElement("p", null,
"Our aim is to disrupt antibiotic resistance by downregulating specifically targeted genes in ",
React.createElement("i", null, "E. coli."),
" We have designed a proof-of-concept project targeting gfp (encoding green fluorescent protein, GFP) and rfp (encoding red fluorescent protein, RFP). These genes are repressed or knocked out by conjugatively delivered CRISPR/dCas9- or CRISPR/Cas9-systems, respectively (see info-box below for the CRISPR/Cas-terms used). These systems were designed with the health of the human microbiome in mind, in order to not disturb its natural ecology ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.sciencedirect.com/science/article/pii/S1021949819300122", target: "_blank" }, "Ding, R-x. et al. (2019). Revisit gut microbiota and its impact on human health and disease. J of Food and Drug Analysis, 3, 623-631. "))),
". With this, we show the potential to successfully downregulate genes encoding OXA-48. "),
React.createElement("div", { className: "info-box" },
React.createElement("p", { className: "info-header" }, "Relevant CRISPR/Cas-terms used by us to ease understanding throughout this wiki:"),
React.createElement("ul", null,
React.createElement("li", null,
React.createElement("p", { className: "text-header font-bold" }, "CRISPR/Cas9-system:"),
React.createElement("p", null,
"This term is used whenever we are referring to the system containing the protein Cas9. When speaking of Cas9\u2019s effect on DNA, the term ",
React.createElement("span", { className: "font-bold" }, "knockout"),
" is used.")),
React.createElement("li", null,
React.createElement("p", { className: "text-header font-bold" }, "CRISPR/dCas9-system:"),
React.createElement("p", null,
"This term is used whenever we are referring to the system containing the catalytically inactive protein dCas9. When speaking of dCas9\u2019s effect on DNA, the term ",
React.createElement("span", { className: "font-bold" }, "repress"),
" is used.")),
React.createElement("li", null,
React.createElement("p", { className: "text-header font-bold" }, "CRISPR/(d)Cas9-system: "),
React.createElement("p", null,
"This term is used whenever we are referring to both the CRISPR/Cas9-system and the CRISPR/dCas9-system, interchangeably. When speaking of their effect on DNA, the term ",
React.createElement("span", { className: "font-bold" }, " downregulate"),
" is used.")))))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.ProjectOverview = ProjectOverview; //# sourceMappingURL=ProjectOverview.js.map } // default/src/components/Content/Lab_Process_And_Parts/LabProcessOverview.jsx $fsx.f[202] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const BulgeWidget_1 = $fsx.r(190); class LabProcessOverview extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "labProcessOverview" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "Parts & Procedures", img: "
", fromColor: "#1D3C4D", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CThe way to get started is to quit talking and begin doing\u201D "),
React.createElement("p", { className: "quotemaster" }, "- Walt Disney, American animation entrepreneur"),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null, "In this chapter, you will find all the information about our work in the laboratory. Our collection of parts, SOPs, procedures, as well as a weekly notebook covering our work is available and should be sufficient information for future iGEM teams to replicate our experiments. Furthermore, you will find information about the safety and security procedures which we complied with during our work. "))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.LabProcessOverview = LabProcessOverview; //# sourceMappingURL=LabProcessOverview.js.map } // default/src/components/Content/Human_Practices/HumanPracticesOverview.jsx $fsx.f[204] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const BulgeWidget_1 = $fsx.r(190); class HumanPracticesOverview extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(203);
});
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "References",
itemTitleColor: "text-blue-800",
items: [
{
title: "Project & Inspiration References",
content: `
- <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4967260/" target="_blank" class="references">Jasovský, D et al. (2016). Antimicrobial resistance - a threat to the world’s sustainable development. Ups J Med Sci, 121(3), 159-164.</a>
- <a href="https://amr-review.org/sites/default/files/AMR%20Review%20Paper%20-%20Tackling%20a%20crisis%20for%20the%20health%20and%20wealth%20of%20nations_1.pdf?fbclid=IwAR1aor_fszMsgDxV8cOaTmFtj59D0OAUqcbDJcf0XGOyBPVxaLJVNcryEMM" target="_blank" class="references">O’Neill, J. (2014). Antimicrobial Resistance: Tackling a crisis for the health and wealth of nations. Wellcome Trust.</a>
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "humanPracticesOverview" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "Practices", img: "
", fromColor: "#1D3C4D", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content" },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CHow wonderful it is that nobody need wait a single moment before starting to improve the world.\u201D "),
React.createElement("p", { className: "quotemaster" }, "- Anne Frank, Holocaust-victim and diarist"),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null,
"Welcome to our Human Practices! This is where we will tell you all about our work beyond the laboratory. Our project relates directly to the Sustainable Development Goals (SDGs) set by the United Nations. This is evident from the 2016 commentary in Uppsala Journal of Medical Sciences by Du\u0161an Jasovsk\u00FD et al. ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4967260/", target: "_blank" }, "Jasovsk\u00FD, D et al. (2016). Antimicrobial resistance - a threat to the world\u2019s sustainable development. Ups J Med Sci, 121(3), 159-164. "))),
", which highlights how antimicrobial resistance could hinder the achievement of the SDGs. "),
React.createElement("p", null, "Conjugaid focuses specifically on the following SDGs:"),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "\nhttps://static.igem.org/mediawiki/2019/2/2d/T--SDU-Denmark--sdg3_inverted.png\n" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("p", { className: "font-black text-lg" }, "SDG No. 3: Good health and well-being"),
React.createElement("p", { className: "italic" }, "Ensure healthy lives and promote well-being for all at all ages"),
React.createElement("p", null,
"Antimicrobial resistance will be the leading cause of death in 2050 according to the O\u2019Neill report of 2014 ",
React.createElement("span", { className: "citation" },
React.createElement("span", { className: "citationtext" },
React.createElement("a", { href: "https://amr-review.org/Publications.html", target: "_blank" }, "O\u2019Neill, J. (2014). Antimicrobial Resistance: Tackling a crisis for the health and wealth of nations. Wellcome Trust."))),
". It is therefore an imperative place to look when ensuring good health and well-being for all."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "\nhttps://static.igem.org/mediawiki/2019/d/d4/T--SDU-Denmark--sdg4_inverted.png\n" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("p", { className: "font-black text-lg" }, "SDG No. 4: Quality Education"),
React.createElement("p", { className: "italic" }, "Ensure inclusive equitable quality education and promote lifelong learning opportunities for all"),
React.createElement("p", null, "Sustainability must be included in curricula all over the world in order to ensure this development. With our educational material, we have engaged students in the conversation around antibiotic usage and biotechnology in a global perspective."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "\nhttps://static.igem.org/mediawiki/2019/6/61/T--SDU-Denmark--sdg12_inverted.png\n" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("p", { className: "font-black text-lg" }, "SDG No. 12: Responsible Consumption and Production"),
React.createElement("p", { className: "italic" }, "Ensure sustainable consumption and production patterns"),
React.createElement("p", null, "According to Jasovsk\u00FD et al, antibiotics must be looked upon as a scarce and potentially non-renewable resource, in which novel innovation is needed. Our project attempts innovation in this regard by implementing novel technologies in our solution."))),
React.createElement("div", { className: "flex-container" },
React.createElement("div", { className: "flex-item logo-item" },
React.createElement("img", { src: "\nhttps://static.igem.org/mediawiki/2019/8/85/T--SDU-Denmark--sdg17_inverted.png\n" })),
React.createElement("div", { className: "flex-item text-item" },
React.createElement("p", { className: "font-black text-lg" }, "SDG No. 17: Partnerships for the Goals"),
React.createElement("p", { className: "italic" }, "Strengthen the means of implementation and revitalize the global partnership for sustainable development."),
React.createElement("p", null, "Antimicrobial resistance is a multi-faceted issue, and needs to be solved by many actors. Thus we have included inputs from both industry and healthcare workers into our project design."))),
React.createElement("p", null,
"In Human Practices, we have considered the ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Human_Practices", target: "_blank" }, "bioethical implications"),
" of working with synthetic biology and antibiotic resistance in order to promote healthy debates with members of society. Amongst other things, we have engaged in dialogue with the general public as well as local politicians of our municipality and Members of Parliament."),
React.createElement("p", null, "In Integrated Human Practices, we reached out to local Doctors Ulrik Justesen and Torben Barington as well as Danish CRISPR-company SNIPR Biome to discuss our project design. "),
React.createElement("p", null,
"In Education and Public Engagement, we have created an open-source lesson-plan for high school students majoring in biotechnology. It consists of educational material (an electronic book), teaching manuals and laboratory protocols to educate within synthetic biology and antibiotic resistance. The lesson-plan was tested by local students as well as their teachers, and will be presented ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Public_Engagement", target: "_blank" }, "later in our wiki"),
". Furthermore, we have hosted workshops for Danish high school students across the country, participated in The People's Democratic Festival, debated with representatives from Pfizer Inc.\u2019s management, the EU Commissioner for Competition as well as several Members of Parliament, and we have participated in many other outreach events. Keep reading to learn more about our engagement with society."))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.HumanPracticesOverview = HumanPracticesOverview; //# sourceMappingURL=HumanPracticesOverview.js.map } // default/src/components/Content/People_Involved/PeopleInvolvedOverview.jsx $fsx.f[205] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const BulgeWidget_1 = $fsx.r(190); class PeopleInvolvedOverview extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "peopleInvolved" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "People Involved", img: "
", fromColor: "#1D3C4D", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content " },
React.createElement("div", { className: "row" },
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CIndividually, we are one drop. Together, we are an ocean.\u201D"),
React.createElement("p", { className: "quotemaster" }, "- Ryunosuke Satoro, Japanese author"),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null,
"Something extraordinary happens when teamwork is done correctly. As Satoro writes, a team is like an ocean; constantly undergoing ups and downs, yet capable of achieving great things as a result of all of the powerful forces affecting it. ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Team", target: "_blank" }, "Our team"),
" has indeed been affected by many great forces, and without them, Conjugaid would never have seen the light of day. Please take a moment to appreciate our ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Team", target: "_blank" }, " team supervisors"),
", who were always there to save us when the seas got rough. Through ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Attributions", target: "_blank" }, "external attributions"),
" we have been able to take our journey with Conjugaid even further, and by ",
React.createElement("a", { href: "https://2019.igem.org/Team:SDU-Denmark/Collaborations", target: "_blank" }, "collaborating with various iGEM teams"),
" we have broadened our knowledge and had so much fun along the way. Our sponsors have provided us with everything we needed to get Conjugaid going and to make sure we did not drown along the way."))),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.PeopleInvolvedOverview = PeopleInvolvedOverview; //# sourceMappingURL=PeopleInvolvedOverview.js.map } // default/src/components/Content/Introduction/Introduction.jsx $fsx.f[207] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const tslib_1 = $fsx.r(8); const React = $fsx.r(3); const Buttons_1 = $fsx.r(104); const BulgeWidget_1 = $fsx.r(190); class Introduction extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
}
UNSAFE_componentWillMount() {
return tslib_1.__awaiter(this, void 0, void 0, function* () {
$fsx.r(206);
});
}
render() {
let accordionData = {
backgroundColor: "bg-blue-800",
title: "Achievements",
itemTitleColor: "text-blue-800",
items: [
{
title: "Bronze",
content: `
- Registration and Giant Jamboree Attendance ✓
- Competition Deliverables: Convince the judges that you have completed the following Competition Deliverables:
#1 Wiki ✓
#2 Poster ✓
#3 Presentation ✓
#4 Judging Form ✓
- Attributions: Convince the judges that you have completed Competition Deliverable #5 <a>Attributions </a>. ✓
- Project Inspiration and Description: Convince the judges that you have completed Competition Deliverable #8 <a> Project Inspiration and Description</a>. ✓
- Characterization: Convince the judges that you have added quantitative experimental characterization data to an existing Part from the Registry of Standard Biological Parts. ✓
Our team has characterized the following part: <a href="http://parts.igem.org/Part:BBa_J01003" target="_blank">BBa_J01003.</a>
},
{
title: "Silver",
content: `
- <a>Validated Part</a>: Convince the judges that at least one new BioBrick Part of your own design that is related to your project works as expected. ✓
Our team has created the <a> part XXX </a>.
- <a>Collaboration</a>: Convince the judges you have significantly worked with one (or more) currently registered 2019 iGEM team(s) in a meaningful way. ✓
Throughout our project, our team has worked with the following iGEM teams: DTU Biobuilders, UCopenhagen, Amazonas Brazil, Lund, TAS_Taipei, Costa Rica & Tuebingen.
- <a>Human Practices</a>: Convince the judges you have thought carefully and creatively about whether your work is responsible and good for the world. ✓
Our team has created and tested a lesson-plan for high school students. Furthermore, we have engaged the public as well as legislators in regards to synthetic biology and antibiotic resistance.
},
{
title: "Gold",
content: `
- <a>Integrated Human Practices</a>: Expand on your silver medal activity by demonstrating how you have integrated the investigated issues into the purpose, design, and/or execution of your project. ✓
We have continuously altered our project purpose, design and execution after discussing our ideas with professor in clinical microbiology at our local hospital Ulrik S. Justesen and with SNIPR Biome.
- <a> Model Your Project</a>: Convince the judges that your project's design and/or implementation is based on insight you have gained from modeling. ✓
Through modelling the conjugation efficiency of our project, we have gained valuable knowledge about our project design.
- <a>Demonstration of Your Work </a>: Convince the judges that your engineered system works.
Based on our results within our two systems (conjugation and CRISPR/Cas), both separately and combined, we have determined our engineered system as successful.
}
]
};
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "introduction" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "Introduction", img: "
", fromColor: "#1D3C4D", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content pb-24" },
React.createElement("div", { className: " row" },
React.createElement("div", { className: "quote" },
React.createElement("div", { className: "whiter-line" }, " "),
React.createElement("p", { className: "text" }, "\u201CWhat is not started today is never finished tomorrow.\u201D "),
React.createElement("p", { className: "quotemaster" }, "- Johann Wolfgang von Goethe, German writer and scientist "),
React.createElement("div", { className: "whiter-line" }, " ")),
React.createElement("p", null)),
React.createElement("p", null, "We are the 2019 iGEM team from University of Southern Denmark (SDU). Our story begins with the meeting of 11 different people. Together, we set out to push the boundaries for synthetic biology by creating a new, innovative solution to the global issue of antibiotic resistance. After brainstorming project ideas, we realized that our personal differences are in fact our biggest strength. By combining the unique skills of each team member, we came up with Conjugaid; a conjugatively delivered CRISPR/Cas-system able to sensitize bacteria to antibiotics. You will learn much more about Conjugaid later on."),
React.createElement("img", { id: "intro-img", className: "my-10 ", src: "
" }),
React.createElement("p", null, "We are very excited to invite you on a journey through our iGEM project! Just click \u2018Next\u2019 and you will be guided on a tour telling you everything you need to know about Conjugaid (you may also use the menu-bar above).")),
React.createElement(Buttons_1.NextStationButton, { nextStation: this.props.nextStation }))));
}
} exports.Introduction = Introduction; //# sourceMappingURL=Introduction.js.map } // default/src/components/Content/FinalWords/FinalWords.jsx $fsx.f[208] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const React = $fsx.r(3); const BulgeWidget_1 = $fsx.r(190); const nuka_carousel_1 = $fsx.r(168); class FinalWords extends React.Component {
constructor(props) {
super(props);
window.document.title = `${props.title} | SDU-Denmark`;
this.carousel = React.createRef();
this.state = {
carouselHeight: 0
};
}
componentDidMount() {
let carousel = this.carousel.current;
if (carousel) {
let constant = 0.5625;
let parent = carousel.frame.parentElement;
let parentRect = parent.getBoundingClientRect();
let newheight = parentRect.width * constant;
this.setState({
carouselHeight: newheight
});
}
}
render() {
return (React.createElement(React.Fragment, null,
React.createElement("div", { id: "finalWords" },
React.createElement(BulgeWidget_1.BulgeWidget, { title: "Final Words", img: "
", fromColor: "#D0E8E6", toColor: "#DCEEF2" }),
React.createElement("div", { className: "content mt-32" },
React.createElement("div", { className: "row" },
React.createElement("p", null, "You have reached the final station - and so our story ends. Thank you so much for your time! We really hope you liked getting to know Conjugaid as a project and as a team throughout our wiki. We have gathered some of our favorite pictures from our journey. We hope you will look at them and feel just as happy as we were in those moments - we have truly had a great time with iGEM, and hopefully that shines through."),
React.createElement(nuka_carousel_1.default, { ref: this.carousel, dragging: true, style: {
height: this.state.carouselHeight,
} },
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" }),
React.createElement("img", { src: "
" })))))));
}
} exports.FinalWords = FinalWords; //# sourceMappingURL=FinalWords.js.map } // default/src/components/application/timelines/index.js $fsx.f[211] = (module, exports) => { "use strict"; Object.defineProperty(exports, "__esModule", { value: true }); const gsap_1 = $fsx.r(91); const getDefaultTimeline = (node, delay) => {
const timeline = new gsap_1.TimelineMax({ paused: true });
timeline
.from(node, 0.3, { display: "none", autoAlpha: 0, delay, ease: gsap_1.Power1.easeIn })
.call(() => {
document.body.classList.remove("rodal-open");
});
return timeline;
}; const getHomeTimeline = (node, delay) => {
const timeline = new gsap_1.TimelineMax({ paused: true });
return timeline;
}; exports.play = (pathname, node, appears) => {
const delay = appears ? 0 : 0.5;
let timeline;
if (pathname === "/")
timeline = getHomeTimeline(node, delay);
else
timeline = getDefaultTimeline(node, delay);
requestAnimationFrame(() => timeline.play());
}; exports.exit = (node) => {
const timeline = new gsap_1.TimelineMax({ paused: true });
timeline.to(node, 0.15, { autoAlpha: 0, ease: gsap_1.Power1.easeOut });
timeline.play();
}; //# sourceMappingURL=index.js.map } // Importing a single entry $fsx.r(217);
//# sourceMappingURL=724868fa-app.js.map