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over its applications in Synthetic Biology like modular cloning to the in-depth | over its applications in Synthetic Biology like modular cloning to the in-depth | ||
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During this planning step we realized, that some of the teams that are already working | During this planning step we realized, that some of the teams that are already working | ||
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Revision as of 15:35, 8 December 2019
C O L L A B O R A T I O N S
Through this iGEM year we interacted with many iGEM Teams all around the world. It was a pleasure working with all the different teams and learn from each other. We got many impressions about other projects and we hope that we could help with our collaborations. We would like to thank every team that collaborated with us!
C O L O N Y
P I C T U R E S
The Colony Picture Collaboration
In our colony picking collaboration we asked other iGEM teams to supply us with data for our picking algorithm. Thanks to this group effort we could build our colony picking unit.
Many repetitive tasks in the lab cost time such as pipetting or picking colonies from agar plates and could technically be performed by robots. However, most commercially available Solutions are simply too expensive and therefore unavailable for manny Labs and iGEM Teams. So one of our goals was to build a reasonable colony picking robot to outsource the tiring task of colony picking by hand.
We used an Opentrons OT-2, the cheapest available pipetting Robot and constructed modules and scripts, that allow other iGEM Teams to turn their own OT-2 into a colony picking robot as well. See here how to transfer your OT2 into a Colony Picker.
We chose to build an Artificial Intelligence (AI) to automatically recognise colonies. While a “hard-coded” solution would be easier to achieve, they lack accuracy and flexibility, key attributes of Synbio projects. The drawback for an AI approach is the amount of data it needs for precise and robust training. With the help of the iGEM-community we thought to overcome this problem. We asked other iGEM Teams if they could provide us with labeled pictures of their agar plates with E. coli colonies, which serve as the basis for our AI. For that purpose we set up a Website.
To participate all that was needed were agar plates with E. coli colonies, a screen and a camera. Other criteria were evenly distributed, distinguishable colonies. The first step was taking the picture. For this purpose, the screen had to be set to the highest level of brightness and display a white image. When you shoot a picture of a common screen, pattern on the picture could appear, so called moire pattern. To avoid this, we supplied each team with diffusion paper, which they can use to cover their screen. A detailed summary is found here.
Following this, the images taken had to be labeled using a web-based tool. Afterwards, the single colonies were marked with a rectangle. In total 250 colony pictures were added to our data set, vastly increasing the performance of our AI in detecting colonies. As an incentive to participate in this collaboration we gave away prizes to the teams that sent us their pictures.
As an incentive to participate in this Collaboration we gave away prizes. The five best teams that sent us pictures got an E. coli plushie and the three teams that sent us the most pictures got a bigger surprise. Thanks to Doulix, we provided these three Teams with an engraved trophy out of glass and vouchers.
Team BOKU-Vienna was able to send us one picture, Team GO Paris-Saclay provided 3 pictures and Team TUDelft could help us with 14 colonie pictures. Team TU Darmstadt could send us a number of 23 pictures, but were beaten by Team IISER Bhopal which send us an impressive amount of 99 pictures.
We thank all the teams for sending us labeled colony pictures. Without your help we would not have been able to train our AI and to let our opentrons OT-2 pick colonies.
T H EG O L D E NG A T E
C O L L A B O R A T I O N
The Golden Gate collaboration
Our team made itself the goal to introduce other iGEM Teams to Golden Gate. Therefore we hosted a webinar, build up a Communication Platform and made an Interlab.
Earlier this year the iGEM headquarters announced changes in their part submission system, one of
which made us extremely happy: BioBricks are now also accepted in Type IIS standard! In our opinion,
this is a huge step, as cloning techniques based on Type IIS restriction enzymes are becoming more
and more popular in the scientific community due to their numerous advantages over classical cloning
methods.
We noticed that more teams converted to MoClo systems in their projects this year but we still think
there is a lot that needs to be done for the iGEM community to really embrace this change. In order
to help teams with the transition into this new system, we had an idea:
What if we could help other teams understand what Type IIS enzymes really are and how they work? If
we could share our passion for Modular Cloning and the value of the PhytoBrick standard? If we could
introduce them to our Marburg Collection and help removing some of the burdens that come with a
switch to a new cloning system?
Therefore we hosted a webinar about Golden Gate, build up a communication platform to give iGEM
Teams a chance to troubleshoot their methods and made an Interlab study. Here we evaluated the
consistency of Golden Gate Assembly protocols across iGEM teams as well as to give teams the
opportunity to gain experience with the assembly method.
We wanted to reach as many teams as possible with our webinar on Golden Gate cloning,
Type IIS enzymes, their benefits and how to use all of this in their own projects. We
started planning everything that we thought should be in the webinar, from the
biological function of the Type IIS enzymes and how they differ from Type II enzymes,
over its applications in Synthetic Biology like modular cloning to the in-depth
strategies when designing your own parts and actually building them in the lab.
During this planning step we realized, that some of the teams that are already working
with Golden Gate might not benefit as much from an entry level webinar. Consequently, we
thought about integrating more complex design questions that we could share with the
community. We finished our webinars with a practical part, showing the part-design in
Geneious Prime – the sequence analysis software we used throughout the year. Firstly, we
explained the different levels that can be built with Golden Gate and then introduced
some more complex designs that are also possible with Golden Gate cloning, such as the
construction of whole operons or space holder parts. Through this we hoped to also reach
teams that use Golden Gate themselves, showing them the possibilities this cloning
system offers.
To reach as many teams on the globe as possible we set up webinars on three different times to cover as many time zones within a comfortable time frame – hoping that this will allow more iGEM teams to participate. The one and a half hour webinars were held on July 30th 10am CET, July 31st 6pm CET and the last time on 8th August 12pm CET by our instructor René Inckemann with the support of three of our team members. We were happy that in total 18 teams participated, as well as some students from the universities of Marburg and Gießen and even a few fellow scientists from the Max-Planck-Institute for terrestrial microbiology. In total we counted around 70 participants. We were also really happy to see Sota Hirano from Doulix on our webinar, who gave us some awesome feedback, stating that he did learn new things about Golden Gate which he was not aware of before!
If you would like to learn about Golden Gate assembly please feel free to watch our webinar through this Link:
We thank all the following teams for participating in this awesome event:
Although we were quite pleased with the result of our webinar, we thought about how to
further connect this Golden Gate community, so we created a Slack workspace for everyone
that is working with this method, enabling us all to exchange protocols and ideas or
troubleshoot issues that might arise. So far there are close to 50 members, but we hope
to find more enthusiasts willing to share their knowledge!
All files used for the webinar are openly available. You can find our protocols,
handouts, slides as well as the files we used for the practical part in Geneious to
construct parts in-silico in our shared Google Drive. Through this we hope to make it
easier for other teams willing to start with Golden Gate cloning, as they can start
designing their first own in-silico parts with our guidance.
The main documents of the Golgen Gate Webinar
→ Download: Golden
Gate webinar slide 2019
→ Download: Handout
Golden Gate webinar
→ Download: Golden
Gate protocoll
To try out the described in-silico cloning you can download the needed sequences as
geneious file here.
→ Download: Sequences
Webinar
As we are constantly working on improving the efficiency of Golden Gate. We clearly
saw the main bottleneck of the reaction was the long assembly time in the cycler.
In the webinar and in ourGolden Gate community weprovided protocols with three different
cycler settings for different applications.
Although these threeprotocols were tested before, they were never compared to each other in
their efficiency.
We conducted our own Interlab study in collaboration with the iGEM Teams from
TUDelft ,
HUmboldt Berlin
and
TU Kaiserslautern
In order to find the best protocol, we supplied the teams with specific Level 0
parts,that should be assembled to a Level 1 vector using the three different cycler
conditions. The provided parts areshown in the table
| 5' Connector | BBa_K2560075 / |
| Promoter | J23119 BBa_K2560031 |
| RBS | B0034 BBa_K2560008 |
| CDS | sfGFP BBa_K2560042 |
| Terminator | B0015 BBa_K2560035 |
| 3' Connector | BBa_K2560080 |
| ORI: | ColE1 BBa_K2560036 |
| Antibiotic-Resistance-Cassette | Kan-RFP BBa_K2560056 |
The Golden Gate reactions should be pipetted together using the given parts and then
be transformed in E. coli and be plated with similar amounts on agar plates. A colony with a
correct assembled
vector would have a functioning transcription unit which expresses GFP, which can be clearly
distinguished from RFP expressing colonies which only carries the antibiotic cassette and
false non fluorescent
colonies.
The ratio of green to non-green colonies was used to evaluate which of the three protocols
yielded in the
highest transformation efficiency and the highest rate of positive clones.
The handout with the used protocols can be downloaded here .
Results
First we wanted to know which protocol worked best for every team. For this we analysed the data of the correct assembled green colonies for every protocol. The data of the different teams are given in table 1 and are shown in as chart in Figure 2. The total colony count is hard to compare directly, cause the numbers differ very much between the teams. This could come through the different transformation efficiencies for example because of the variating competent cells the teams used.
The results in Figure 2 show, that the protocols work differently for each teams. In generell the most colonies were
received with the long Troubleshoot protocol, but also the Rapid and Improved protocols show high numbers of correct colonies.
But which protocol is the most efficient one?
To compare the results of the teams the ratio of the gained correct colonies between the
protocols were averaged. The data are shown in Figure 3 and show, that the long Troubleshoot
assembly works the best. The fastest assembly (rapid) works the second best. This is very
interesting, because this assembly takes significant less time.
We would propose to take the troubleshoot protocol for complicated assemblies and the rapid
protocol for easy assemblies like lvl0.
B E R L I N
Berlin Collaboration
iGEM Berlin engineered the freshwater algae Chlamydomonas reinhardtii to degrade
Polyethylene
terephthalate
(PET). For this goal they also built a bioreactor for their chassis. We sent
them our UTEX
strain to test if
their photobioreactor is capable of cultivating other phototrophic organisms as
well. They gained
additional
experiences on growth conditions for phototrophic organisms and we could see our
strains grow in
a bioreactor environment.
Click here to learn more about there project Chlamylicious
D E L F T
Delft Collaboration
Delft Collaborations
iGEM Delft created an all species encompassing toolbox, based on orthogonal gene
expression and
host
independent plasmid replication. We exchanged multiple times over Molecular
Cloning and toolbox
designs.
Additionally, we sent them parts in the PhytoBricks standard.
Click here to learn more about there project
project SCI-PHI 29
U CD A V I S
UC Davis collaboration
UC Davis iGEM team is one of the winners of the Opentrons competition in 2019. iGEM Marburg won one OT-2 during last year’s edition of the competition and got contacted for a collaboration by iGEM UC Davis along with other winners from 2018 and 2019. The goal for this automation InterLab collaboration was to collect data to determine the accuracy and reproducibility of the OT-2 compared to humans. To do this, UC Davis proposed to perform the standard iGEM protocol on serial dilutions with silica beads and fluorescein, each manually and with an OT-2. In total iGEM Marburg did four dilutions; twice with an OT-2 and twice manually. The data was then sent to UC Davis via the spreadsheet designed by them. Finally, iGEM Marburg aided by creating a Slack channel for all iGEM teams with an OT-2 to facilitate better communication and exchange in the future.
In total, seven teams participated and sent their data to UC Davis which are
shown in the
figure below.
According to UC
Davis, the fluorescence data was standardized using the iGEM MEFL conversion to
standardize
the plate reader
measurements. Subsequently, the data was fitted using a linear regression model,
whose
quality is quantified
using the
r2 metric (it ranges from 0 to 1 with higher value indicates a better fit).
For the fluorescence the r2 value from the OT2 data beats human r2 value with
0.94 and 0.90,
respectively.
Interestingly, for the silica beads, manual measurements yielded a more
satisfactory result
with r2 value of
0.90
compared to 0.82 from OT2 measurements. One possible explanation could be the
speed of the
OT-2: it took
twice as long
to complete this task with the OT-2 compared to a manual trial. This could lead
to the
sedimentation of the
silica beads
as the OT-2 went through their runs; therefore worsening its results. Also, as
shown in Fig.
2b, one team’s
result
deviates a lot from the rest of the teams, maybe if their result is taken from
the overall
evaluation a more
comparable
result between OT-2 and human can be observed.
More information about this collaboration such as individual team data can be found on UC Davis wiki: Wiki UC Davis Collaboration
U F L O R I D A
UFlorida
This semester we had the pleasure of mentoring UFlorida for their Wiki
design. Our member, Rene Inckemann, aided UFlorida in their quest to create
a visually accessible Wiki, similar to what we did this past iGEM season.
We helped shape UFlorida’s website into an interactive and appealing
landscape, while maintaining the integrity of their goal and project. We
advised them on their font choices- specifically their use of bold and
italicized words-, along with answering questions about our work with the
visually disabled community and utilizing our “Accessible Wiki Guide” from
the previous year.
Besides answering questions specifically about their project, we also
offered valuable information about when to start working on the Wiki and
questions about coding. Working with a young team like UFlorida offered us
a great mentorship opportunity.
I N T E R A C T I O N S
W I T H
O T H E RT E A M S
Interactions with other teams
Mike the microbe
We participated in the “Mike the microbe” collaboration of the iGEM Team US AFRL CarrollHS. They sent us their virtual mascots Mike the Microbe and Chia the Chitinase so we could take pictures with them to connect all iGEM teams around the world via social media. It brought us great joy to join to this collaboration and let the two little buddies visit our daily life in our incubators for cyanobacteria.
Postcards Düsseldorf
iGEM Düsseldorf called for their traditional collaboration project, a long year postcard campaign with the purpose to educate and to show the public about the different topics and projects in Synthetic Biology and especially regarding all the iGEM projects. With interesting designs and a short project abstract iGEM teams could spread their ideas using the postcards to share their thinking to each other and the society. Our title and postcard design was about our photosynthetic project „Syntex“. You can see Synechococcus elongatus UTEX 2973 direly needing a refreshing drink, a cute little draft from our team member Jonas.
iGEMxSDGs challenge
IGEM Costa Rica, Tuebingen, and TAS Taipei created an iGEMxSDGs challenge as collaboration. They asked us to choose four of the Sustainable Development Goals (SDGs) proposed by the United Nations (UN). We joined this collaboration challenge and spread it on our social media account. We decided on the following goals:
Who wore it best? #labcoatchallenge
The Stony Brook iGEM Team challenged us to design an outfit for a runway from a standard lab coat. The outfit had to be created by tying, not using duct tape, cut or sew, etc. After designing our outfits four of our members presented their style on our ‘runway’ while paparazzi took their photos. As an outfit we chose a dress highlighting the neckline, a shoulder free dress, a knotted dress and a skirt to bring glamour to the daily routine in the labs. Especially our guys were rocking their outfits as you can see here
M E E T U P S
Meet ups
iGEM Spring Festival in Bonn
From the 3rd to the 5th Mai we met other iGEMers at the Spring Festival in Bonn. Professors
of the university of Bonn presented their research fields which was met with huge interest. We heard
lectures
about immunology and neurosciences, as well as reviving mammoths. During the lectures,
specific
topics were deepened in small seminar groups, such as the ethical tenability of CRISPR
applied
at humans and other animals. At the poster sessions we got the first time in contact with
other teams’ projects. Afterwards, we had the opportunity to vote for the best project,
which
iGEM Düsseldorf won. Besides that, the meetup was filled with a lot of awarding
categories.
We
competed in the categories for “Best Video”, “Best Poster”, “Best Project” and a single
person
had the chance to win the award for “Best Meme”. The iGEM Team Bonn offered spontaneously
a
little football table tournament, where two of our team members won. In the evening we
learnt a
lot about Bonn as a city from a guide living there for ages. Subsequently, the iGEM Teams
met
in
several groups in restaurants and bars. We merged with iGEM Kaiserslautern and exchanged
our
projects and ideas for the rest of the evening.
On the second day, the iGEM Team Bonn offered a competition out of three rounds, where we
competed against other teams. We were asked iGEM specific questions, had to solve riddles
for an
escape room and finished a lab challenge afterwards. Our team could compete until the end
and
won the prize! Thanks again to Doulix for this awesome prize!
We particularly thank the iGEM Team Bonn for the huge effort and the great meetup!
Erlangen collaboration meetup
On the 17th till the 19th of May we attended the collaboration meetup hosted by the iGEM Team Erlangen 2019. There, we met the iGEM teams of Straßbourg, Tübingen, Vienna and of course Erlangen. Apart from listening to the interesting talks from the various professors there, we made a lot of new friends and took the chance to talk about possible collaborations with the other teams and also promote our collaborations, such as the colony picking project and the Golden Gate webinar. We would like to thank iGEM Team Erlangen very much for organizing this wonderful meetup and are happy to see them soon in Boston.
Düsseldorf meetup
From the 5th till the 7th July we went to the HHU for the German meetup. Many lectures with interesting topics
were offered by the Cluster of Exellence of Plant Science (CEPLAS)by Prof. Andreas Weber
but also non-scientific aspects like coureses on presentations. We tried to visit all of
them
while presenting our poster in the hall. There we got in touch with other iGEM Teams and also
learnt
about their projects. We promoted our Webinar project and laid the foundation stone for
future
collaborations. On the second day, iGEM Düsseldorf organized a panel discussion about the
issues
concerning green genetic engineering. Thereby, we got great input on how to structure our
panel
discussion and which subjects should be mentioned. Beside the regular program organized by
iGEM
Düsseldorf we used a lot of chances to get in contact with other teams. After the
presentations
a couple of tourist guides showed us the beautiful old town of Düsseldorf and explained
some
interesting historical facts. Afterwards we had again the possibility to meet with other
iGEMers
in a pub and talk about the projects, especially to find weaknesses or opportunities for
collaborations. This really went successfully due to the time we spend with iGEM Team Aachen, Kaiserslautern and Hamburg. The iGEM Team Düsseldorf also arranged a game contest between the teams,
where
we lost in the final to our good friends from Potsdam. Congrats!
Thanks again to iGEM Düsseldorf for the huge effort and the great meetup!
Vilnius, Lithuania – Biohackathon Lab App 2019
In August 2019 our Team participated in the Biohackathon Lab App, organized by the
Vilnius-Lithuania Team 2019. Together with the iGEM Teams Stockholm and UCopenhagen, as
well
as
several non-iGEM Teams we sat together for a weekend packed with problems, ideas and neat
solutions.
With the task of creating an application that tackles a self-found problem, we sat
together
and
brainstormed about what could be done.
During this time we were starting to work more with the Opentrons OT-2 and soon realized
that
some
of our biologists had trouble integrating protocols in the robot themselves, as this is
primarily
done by writing the protocol in Python and pushing it directly to the robot – no
convenient
application to change parameters, such as the amount of reactions,was included in the
protocols.
In our heads the idea to have a user-friendly GUI grew more and more and soon we found
ourselves
standing in the Vilnius University Life Sciences Center.
“We” were two biologists and one mathematician, so we talked to some of the other
participants
that did not have a team yet and found the perfect fit to complement our small team: Nour
Alsamsam
was happy to join us for this weekend and provided us with his coding skills – huge thank
you
to
you, Nour!
Together we tried our best to come up with a suitable app and presented it in the end,
having
more
ideas in our heads than we could implement in such a short time – it was clear for us,
that we
would continue with this project later on!
In conclusion, the Biohackathon was an awesome and productive meeting of the iGEM
Teams
Stockholm, Copenhagen, Vilnius and Marburg. We talked about our projects, offered each
other
feedback on current issues in the lab and shared many ideas.
We are very grateful that we got the opportunity to meet so many experts from different
fields,
which really helped getting a fresh perspective on our project and are now looking forward
to
meet
the other teams again at the Giant Jamboree!