Team:USP-Brazil/Human Practices
Welcome to Integrated Human Practices
The _genSwitch project was first thought to be a basic research, however , during the year, as we received many feedbacks and suggestions we noticed that gradually our project turned out to be more of an aplicate research than basic research. At the end of integrated the project had a huge diversity of applications in the industrial market.
These gradual change can also be visualized, when we see the transition of the stakeholders that we met. Changing from Universities clubs and facilities to market. During our integrated, the team also consulted with different specialists.
Why
In order to start the activities related to integrated, in March we decided to present , in March 27, our project in the Synthetic Biology Club at our university (University of São Paulo).
What
This club, nicknamed Symbio, is formed by students, alumni and the inner community interested in subjects about synthetic biology. They reunite once in a week to talk about different subjects in the field, inviting other specialists and members of institutes and industry. We were invited to talk about the genSwitch and what is iGEM. We received a lot of feedback, tips for the competition (from those who had participated in iGEM previous editions) and also tout 2 new members for the team. Most of the feedback was applied in our project afterwards.
How
Before discussing with Symbio, the project was elaborated based only on the construction approach A of the genetic circuit, however in the club it was raised the discussion about how risky it is to develop an project based solely in one plan, since if an problem occur it could lead to the project inviability. The club gave us two suggestions , that we integrated in the project, to solve this situation. The first was to create an approach B (BBa_K3095011) with a genetic circuit simpler than the Approach A. As we are using light as output for our circuit, the second suggestion was to add an analysis involving the light promoter and the chemical promoters, such as the arabinose promoter and LacI. Human practices is one area of iGEM competition. It is really important, since it’s where the teams can see how theirs works affects the world and how the works affect the project. Even though it is really important to develop human practices, our groups were having a problem with determining how to do the integrated human practices, , mainly because our project was a basic research. To solve our problem, we decided to consult Symbio. There we were advised about how to decide the integrated human practices. First we were advised to think about possible problems that our project could solve , since If there are problems, there are also customers interested in finding the solution. Together with Symbio, a brainstorm was done to find these customers. As our project could alternate between two states of gene expression by using blue light as output. Between the suggestion we had the utilization of our system for the insulin production control for diabetics patients and for the production of components that required the combinations of two types of subproduct, we also received some advice to look for projects associated with metabolic pathway. Overall after the brainstorm lead our integrated to interactions with the industry.
Why
In July 8 we gather together with the scientific representant and founder of Biolambda, Caetano Padial Sabino. Biolambda is a biomedical optics company, that focus on development of highly automated and precise light-based technologies for optics and photonics applications.
What
As our project involves the application of LED light during bioproduction, the most part of our wet lab experiments involved testing our engineered bacterias with LED blue light, the one that we chose for our main circuit. We also needed to test our light-inducible plasmids, kindly donated to us by Christopher Voigt's lab from MIT. This way, we contact Caetano to see if he could help us with some solutions in the lab. During the meeting we explained what is iGEM, how USP-SP teams are formed and what is our project about, specially all the lab experiments that we were doing and planning. After the presentation, he suggested to establish a partnership with us, specially because BioLambda area is very correlated and suitable in our experiments. He also kindly lent us the LEDbox, an irradiation device applicable to large samples used in Biophotonics, Photobiology, Photochemistry and Photomedicine research. This equipment has irradiance levels up to 100 mW per square centimeters and warranties homogeneous and precise light transmission among the entire illuminated surface. It's also protected against short-circuits, overheating and burned diodes (life spam over 10,000 hours). Even if LEDbox is available in more than 65 wavelengths in the UV-VIS NIR spectra, we utilize it in only 450nm of the light spectra, because it give us the LED blue light that we needed.
How
In this meeting, we received the Ledbox. The light device that we used for all the _genSwicht experiments along the year. We also had a fruitful discussion about the importance of measuring the exact timing of LED exposure that activates gene expression and isn't toxic to the bacteria at the same time, something that we had to add in our lab schedule. Caetano remind us that led blue light can be utilize as a decontaminant, as it has bactericidal properties especially at the range of 400-450nm, so he suggested that we stimulate the bacterias with 450nm an plus. In this discussion we also learned that LB medium may have a negative effect in our light stimulation because of its. Lastly, this meeting was incredibly productive and we could exchange some lab experience and received a lot of input in our project!
Why
To see the applicability of _genSwitch in bioproduction we visited IPT BioNanomanufatura Nucleus, Laboratory of Chemical Processes and Particle Technology. IPT is an institute of technological research associated with the Economic Development Department of São Paulo state. It is one of the largest research institute in Brazil, having an history of over 100 years assisting the public and private sections with services and technological solutions, in order for business growth and improvement of quality of life. For that IPT has qualified laboratories and teams of highly qualified researchers and technicians, acting in four majors departments: innovation, research and development; technological services; metrological support and development; information and technological education. Inside innovation, research and development we have the BioNanomanufatura Nucleus. A department focused on products and industrial process development , that meets the sustainable, social, economic and environmental principles. To get these solutions, the department has an multidisciplinary profile, having nanotechnology, biotechnology process and micromannufacture ultra-precision metrology.
What
When we arrived at IPT, we first got a guided visit to their laboratories. One section of IPT laboratories were dedicated to biotechnology. Inside it, we saw the bioreactors used in their projects. We were also able to see a section dedicated to cell culture, plants growth , protein analysis, toxicology analysis. While doing the visit, we got to know some of the works developed by the institute. Here we present 3 of them:
1.Plastic made out of sugar cane: Project focused on the development of biodegradable plastics , which takes 6 to 12 months to degrade, as an alternative to the usual plastic , which takes 200 years to degrade.
2.A cost analysis for biotechnological chemical production: Project focused on analysis of performance projection of biobutanol and fumaric acid production, estimating the costs related to those process.
3.Cosmetics based on residues: Project focused on the development of biosurfactants produced by microorganism, as substitute to synthetic surfactant derived from oil.
After the guided visit we did the presentation of our project to the researchers of BioNanomanufatura Nucleus. We discussed with Dra. Natalia Cerize and her team about the applications of our project.
According to then our project _genSwitch have a lot of applications since it’s proposal of creating two memory states of expression capable of switching in response to a single type of output would increase the efficiency and the control of bioproduction process. By efficiency we can say that having two states of memory, could increase the possibilities of production combinations. That could lead to optimisation of lysine production, where it’s production is tightly associated with growth. In lysine case our project could be used as a platform to program two controllable states of expression, one state with expression of genes associated with growth and the other state with lysine production genes, allowing producers to alternate expression states according to their need.
The alcohol production could also benefit from the same system of controllable alternation of growth state and production state. It’s known that high levels of alcohol in bioreactors leads to the death of fermenters, so the use of an controllable system of alternation between growth and production could decrease the chance of death allowing the achievement of an even better optimal state of production.
Until here, we considered states of expression alternating between growth and production, however even in situations where only growth is interesting our genetical circuit could be applicable, since we could promote the expression of genes related to biomass gain, increasing the gain.
Another advantage of our project would be associated with the reduction of product cost by the use of light inductors. Nowadays the bioproduction industry use chemicals inductors, such as IPTG, that are quite expensive. Together with it we also have the cost of removing these inductorns from the final product, which increase the industrial expenses and the product prices. That is the case of insulin, that has an production associated with IPTG inductors. The use of light, in these situation, a cheaper inductor could decrease these spending.
Another interesting advantage of utilising light as inductors would be the velocity of explosion. In some situations, such as production of recombinant proteins the velocity of IPTG pouring defines the quantity of inclusion bodies will be formed, in other words, slowly IPTG exposure leads to fewer inclusion bodys in the final product. As light inductors are faster than chemicals exposure, its use could increase the production.
How
After discussing with IPT, our project received a lot of feedbacks related to applications of our project in bioproduction and we were advised to get in contact with food, healthy, agro and bioproducts industries. We also calculated together with IPT the estimate cost of chemicals compounds in the industry. For IPTG we have that, the cost of each gram is 50$. Normally in a biorreactor of 7 L we spent 100$, if we consider that in the industry 1 bioreactor have 200 L, we have that an industry spends 2900 $ for each biorreactor.
Why
As the project advanced and the visits to biotech industries kept goinr, we realized the diveristy of potential application in bioproduction. With that, we started to think about the paths we could take if we wanted to keep with the project even after iGEM 2019. With that in mind, we talked to Caetano, Biolambda’s CEO. Biolambda has a lot of characteristics in common with what shall be a company based on Genswitch: a startup founded by undergraduate/graduate students with photobiology as its business core.
What
We had an interview with Caetano about the process of creating a company from a wet lab idea and we also discussed about photobiology market. When did you have the idea of making a company based on photobiology devices? Since my first semester in physics graduation I worked in a project related to this. So during these first years I learned about different technologies in this field, mainly focused on bacterial growth control. I always intended to start a business. I noticed that in research laboratories there was none companies in the market that could provide practical and precise equipment to be used in photobiology. I used to build my own experiment equipment for this so I thought: What if I started to manufacture and sell devices for researchers like me who need this kind equipment? What is your main market? Nowadays we work only with equipments for scientific research, mainly in Brazil. Among these devices, the most popular is the cell plate irradiator, which is the one we lend you for your iGEM project. What are the advantages of working with photobiology and optogenetic instead of using the classical methods? The biggest advantage is that it allows us to make very specific changes in a desired target. For example, we can use it in a therapy by combining a certain molecule that interacts with a target and which is also capable to absorb a very specific light frequency. By combining this two properties we have even more control over a molecule for the design of a certain therapy. Another advantage is that it is almost residual-free, following the global environment concern. LEDs need few material to be produced and has a really long lifetime. Are there many laboratories that works with optogenetics photobiology or optogenetic in Brazil? Yes! Actually, Brazil is worldwide leader on some fields reagarding the application biological application of light, mainly in therapeutic applications. In regenerative medicine, used for regenerate wound, treating pain and inflammation, Brazil is very relevant. Another application that our country is protagonist is the use of light for the control of microorganisms. But in optogenetics I haven’t seen a lot of research in Brazil. What would it be this application of control of microorganisms you mentioned earlier? Eliminating pathogens by using light or other methods mediated by light. For example, UV light is capable of eliminating any organism, but there is also photodtnamics, which is a therapy that we ‘ve been working with lately. In this therapy approach, we use a molecule that is able to absorb a certain light and releases toxic molecule in the very specific site where the molecule is and the light encounter each other. Are there industries using photobiology nowadays? Yes, we have some of these in Brazil. Actually the application of usage of light in medicine also includes some common applications, such as tattoo removal and tooth whitening. A lot of industries starting to explore this market. Does Biolamda have any connection with your PhD? Totally! My PhD thesis was about eliminating microorganisms by combining light with medicines that interacts with light. This interaction is known for more than a century, but no one really knows the exactly mechanism that is involved in this process. The damage caused by the molecule can be either at the DNA, in some proteins or in the cell membrane, but nobody knows what is the main target in this mechanism. According to what I’ve seen, it seems like that the main target of this mechanism are proteins. Once we confirm that this is the way the cell dies, we can develop a combined therapy of this medicine with other molecule that attack bacteria and cells by other means. I’m currently studying combining this molecule with another one that targets the bacteria membrane. What is the future for Biolabda? We are currently working with three fronts. The first one is using the light for preventing contamination at industries. The second one is the use of light for treating wounds exposed to infections. The third is the newest is developing a tool to help acne treatment.
How
With the interview with Caetano we learned more about the advantages of photobiology and were able to understand better what is the Brazil stance toward this type of research.
Why
SuperBAC is a biotechnology company that seek to transform challenges into sustainable opportunities. With the principle of self-remediation of nature, they formulate specific blends of microorganisms, enhancing their effects to solve each demand. We consult them by indication during our meeting with IPT bioproduction group. We had a difficult time to send a good email that could interest them, after a failed attempt and some tips from our advisors we were able to pique the interest of the company. Normally, they don't work with OGM but they have an open innovation hub and an innovation center and for that reason they were interest in our project. They have international partnerships in Singapore and Israel.
What
They invited us to visit their plant focused on innovation and technology for all business models of the company at Parana, (PR) but because of the time we couldn't go. We made a skype meeting with SERGIO LUIS MOREIRA, Consultant and Manager of Research, Development and Innovation in Biotechnology, with experience in Industrial Biotechnological Processes, in the areas of Agriculture, Pharmaceutical, Biofuels, Environmental and Consumer Goods. We made a quick presentation on what is iGEM, about our team, a general knowhow of our project and the possibles applications on bioproduction. Since Sergio has a great experience in the area, he challenged us with some questions about our project that we haven't thought before. If you're working with steel tonnels how would you put led light inside it? Why would bioproductions centers change some well established in the literature inductors, such as temperature, for your project? He also said that in the future we could test our system in their bioreactors.
How
After the meeting, we made a group meeting to discuss what we could do to solve this questions. The steel one, we are still thinking of, because it's something that we plan to solve it in the future since is an engineering problem. However, the temperature one we made a research on the possible benefits that genSwitch could provide in that case.
Why
In September 3, we visited Sakata Seed research division, located 1 hour and a half away from our university, outside the city. Sakata seed is an centenary company known for its contribution in varieties of vegetables and flowers that always thinks about their costume well-being. Their production are totally natural without the use of transgenic.
What
Even though they don’t use transgenic, their research team are always keeping up with the emerging innovations. Because of that, our team got the opportunity to present our project, receiving valuable feedback.
How
The discussion with Sakata research division, gave us the overview that our project is in principle a tool, since we can use it in any species. And if its a tool, it can be patented. Regarding the application of the project, we receive suggestions that our system could be used in the alcohol industry, as it was also said in our IPT meeting and could be of service for the agricultural input. After brainstorming we talked about the possibility of _genSwitch application when thinking about fotoperiod, where we could, by light induction promote expression of molecules associated with flowering. One interesting topic that emerged during the discussion was “what is the limit to be considered an transgenic”. If an vegetable is naturally grown but the agricultural input used are produced by an transgenic organism, should we call it natural , or, transgenic? The new tool for DNA editing, CRISPR, is modifying , directly, genes and sequences in organism. Should organism edited by CRISPR be called transgenic? According to our survey of Brazilian legislation, the CTNBIO Normative Resolution No. 16 of January 15 declares that every product edited by techniques of DNA editing that don’t leaves traces of DNA/RNA recombinants known as Precision Breeding Innovation (PBI) must go through a consultation with CTNBio, where it will be defined if the product fits in the legal definition of transgenic. The first decision based on this resolution came in June 2018, involving genetic alterations by CRISPR, in yeast. After passing through an extensive analysis the yeast was declared a non-transgenic by the committee. Even thought yeast edited by CRISPR wasn’t considered an transgenic. These situation and the discussion done with Sakata brought out the necessity of a deeper reflection about these topics, especially if we think about more complicated techniques and situations that could arises as more innovations are made.
Why
We wanted to consult an health pharmaceutical company to see if they could be interested in genSwitch technology and how could we improve it to attend this sector. For that reason we sent emails to the President/General Manager, Rogerio Frabetti; the Head Medical Affairs & Pharmacovigilance, Helio Guy Osmo and the finance director Wegton Nascimento from Zambon. Zambon is an Italian Pharmaceutical company that has been present in Brazil for 60 years, and has a team of approximately 300 employees throughout the country. Zambon was established in 1906 in Vicenza and is headquartered in Milan within OpenZone. The company operates in the pharmaceutical and chemical business and is present in 20 countries with subsidiaries in three different Continents – Europe, America and Asia – with manufacturing units in Italy, Switzerland, France, China and Brazil. Zambon products are commercialized in 84 countries. Today, Zambon's Brazilian branch represents the company's fourth largest operation in the world and has grown considerably in recent years. Their products are focused on Pain, Respiratory tract, Women's Health, Neurology and Gastric Disorders lines.
What
At 09 september we went on a in person meeting with Rogerio Frabetti (President/ General Manager at Zambon), Helio Guy and Wegton Nascimento. We presented our project for them and also explained what was iGEM and what, in general aspects, genSwitch was. They gave us an important feedback on the presentation: first we had to exhibit our previous prizes at iGEM as TEAM USP then, what are the possible applications of genSwitch and give more examples in health, make a cost analysis and seek for a patent in order to interest companies investments. They also explain to us a real company problem and their focus on respiratory tract diseases and asked us if genSwitch could do something about that. Furthermore, they proposed us to participate at Zcube. Zcube, Zambon Research Venture, is the division that actively deals with innovation, scouting for modern health solutions in the field of Life Sciences, supporting the validation of drug delivery systems, medical devices, biomarkers, diagnostics, digital health and big data solutions. Zcube invests and supports innovative ideas from their early development phases right through to the market launch.It is made of three units: Zcare, Open Accelerator and OpenZone. For more information about it just click here
How
After this meeting we changed our main presentation, and added context and visual figures that a total lay person can understand and also understand its importance as a tool. Also we start looking for possible genes that activate interest products on respiratory tract diseases such as asthma. And furthermore, we sent emails to AUSPIN (USP Agency of Innovation) in order to seek a patent information meeting.
Why
After Zambon meeting, which we obtained an opportunity to possibly participate on a startup incubator, we realized that maybe our project could be patentable. So, after reading the iGEM patent guides (made by other teams) and also sending emails to de headquarters, we set up an in person meeting with AUSPIN (USP Agency of Innovation).
What
We talked to a patent specialist from our university, explained our project and the aspects that made it innovating. We wanted to know which steps we had to follow in order to submit a patent in Brazil. In general lines, the laws of patent are different from each country, like in our country that is something called "periodo de graca", a time of 12 months that you have to submit your patent even after presenting your project in some seminar. In Brazil, gene parts are not patentable, neither an organism. Specifically in our case, every innovation made at the university is mandatory to be submitted via AUSPIN, belonging 45% of it to the University and 10% to the agency, although the process is total free. You can see the regulation of our specific case in here. The most important thing that we learn was that in order to the patent evaluator consider it innovating it has to NOT be a logic step putting together all the different kinds of previous works related to the area.
How
This meeting was very important to us because we had many doubts if our project was really innovating or not. Nevertheless, we discuss if we were going to invest our time in writing a patent-like report. After a while trying to review more the literature, we decided that we were not going to apply for a patent because our project was a logic step of some different papers, one of them we link it here. As we experience a hard time trying to know the iGEM rules, we also thought that maybe the headquarters could let this more clear at the general wiki. In the tab "rules of conduct” (or in FAQ) could be added something about this topic, whether is an accepted action or not. And here we link the patent guides made by other teams that can help in the search for some answers.
Why
In October 4, we had a meeting with Bayer Center of Expertise in Tropical Agriculture (CEAT) Director, Dirceu Ferreira Jr. A section dedicated for promoting innovation for tropical agriculture through public–private partnership. In the meeting, we presented what is the iGEM competition and talked about the 2019 project, _genSwicht. It was the first time that an iGEM team contacted the Bayer branch in Brazil, and they were really interested in understanding what was iGEM.
What
In regards to our project, the applicability of our project was numerous, principally when thinking that the genetic system could be later developed in any organism. In the industry nowadays, this system could be really useful in yeast. Also it could have an applicability in bioremediation, by developing an system of soil enrichment for agricultural interests. We also talked about possibilities in the far away future, were we could implement this system , on a large scale, in plants in order to enrich their production of nutrients, or to increase their growth on large plantations in an automated way. A part of the discussion was also dedicated for the bureaucracy involved with intellectual property protection. We were instructed about the importance of having an patent when developing an project that has a innovative idea. We discussed the importance of not revealing totally the project idea before having properly goten the patent , as in some situations it could lead to having the idea stolen by anybody.
How
This meeting was very important for us, since we were able to visualize more applications to our project. We also received one suggestion given by Bayer CEAT. It was to always sign an non disclosure agreement (NDA), before presenting the project to the companies. The NDA contract is an agreement by which the parties agree to the no disclosure of information covered by it. NDAs are normally signed when two companies or individuals consider doing business together and needs to assess the potential of the partnership. There are two types of NDA, the bilateral ,where both parts sign and receive restriction according to what is written in the document, and the unilateral, where only one of the involved sign and are restricted.
Why
Core Us is a scientific consultancy company which works with any company that needs scientific advice about biological subjects. The company consists of academics with experience on industry, and because of that, they are able to successfully bring useful insights from scientific information to executives without any expertise in science. Their solutions range from scientific training for employees of a company to literature reviews for assessment of a new product . To bring these products, it relies on three main expertise: business, communication and science.
What
We had an interview with Gustavo Belchior, CEO of the company, and discussed about the importance of scientific dissemination nowadays and how to introduce an idea such as our project to an audience with no background about it. Q: What are the main difficulties encountered by a startup in Brazil? A: Ricardo Amorim, the top Linkedin influencer in Brazil, once said that the country is one of the places with the most entrepreneurial people. They have an intense entrepreneurship will, but they lack the ambition to build a big company. There are some, such as yellow – a successful bike rental company – but these cases are very rare. A crucial step to attract investments is to show the size of the market, and maybe a lot of startups forget to bring this up. Q: What is the main media for scientific dissemination? And which one is the best one? A: The most popular nowadays is Youtube and videos in general are usually very effective for that. Social medias such as Facebook and Instagram are also very useful. Still, face to face presentation is remains as the best option. An example of that is an national event called Pint of Science, which reaches thousands of non-academic people every year. In the end, videos are the most used platform for this purpose but the best option will be the brings you the best connection to your audience. For older people, newspaper is still the best choice, while social media is the best for the young. Q: Is there any biology filled that the companies need scientific advice? A: It varies between our cliente. Among agriculture companies for example usually are interested in OGMs, genetic editing, pharmacology, plant pests, plant physiology and population dynamics. Meanwhile ,pharmaceutical companies tends to look for answers regarding drugs pharmacokinetics, toxicity and mechanism of action of a drug.
How
The interview with Gustavo, let us have a deeper understanding about what is a Startup and the productive ways for science communication. Since Gustavo has deep experience on presentations of science to various types of audiences, we presented him our project. He gave us some tips on how to make a presentation more dynamic and catchy. Another important tip he gave us was to always show numbers regarding the impact and the market size a the idea may cover. Finally, Gustavo suggested us to look for Mauricio Baptista, a professor that has deep experience on optogenetics and had even submitted a patent.
Why
As one of the components of our project is the led light, specifically the blue one, we wanted to consult a specialist in the area to clear our doubts about the subject and make sure we were going on the right way.
What
We interviewed professor Cristiane Tavolaro. She is graduated in Physics from Pontifical Catholic University of São Paulo (1985) and has Master in Instrumentation on Nuclear Physics from the Pontifical Catholic University of São Paulo (1992). She is a master assistant professor at the Pontifical Catholic University of São Paulo, working in the courses of Physics, Electrical Engineering, Production and Civil, Mathematics (in-person and distance learning) and Computer Science. Has experience in Physics Education, acting on the following subjects: teaching modern physics, physical optics, acoustics and new technologies for teaching physics. She is a founding member of GoPEF - PUC-SP Physics Teaching Research Group, certified by CNPq and founding member of Marcelo Damy Advanced School of PUC-SP. She is coordinator of Engineering and Technology Laboratories at PUC-SP. The interview is written below: Q: Could you characterize led light? A: LED is a semiconductor that conducts electricity when applying ddp, because the electrons go from the conduction band to the valence band and when they come back they emit light at a well-defined frequency. Due to that characteristic exists red, orange, yellow led light. Q: What is the main difference between led and a common lamp? A: Incandescent lamps produce a very interesting spectrum but a good deal of the lamp's power is lost in form of heat, which reduces the lightning capability. This doesn't happen on the led light and also the blue light peak on incandescent lamp spectrum is very low compared to the blue one. Q: Does the blue led, at wavelength 450-470nm, have any particulars effects that come to mind? A: The blue led was one of the colors that came later since there was a difficulty in doping the semiconductor so that the electronic jump was bigger and produced the blue. Q: Is it correct to say that led is safer for human health than other types of light used in lamps? A: No. Blue light interferes with the circadian cycle and it turns out that blue light causes macular degeneration, destroying retinal cells. The industry has not yet provided available and efficient solutions, there are filters in electronic devices (mobile and notebook) but artificial LED lighting has no filter. Q: Is the use of led more economical than other forms of lighting? A: Yes. In 2016, industries were able to produce in one semiconductor crystal the energy bands that produce all colors, with a big peak in blue. The LED is much more cost-effective than other light bulbs.
How
After that interview we realized that our project has an important safety issue regarding the blue led light. However in every experiment that we use the blue led box, we weren't exposed to it. In addition, in a bioproduction scenery the led light would be inside the tonnel and not exposed to the humans. Furthermore, we also are thinking of doing appropriate glasses with blue light filter in order to manipulate the biorreactor, but for a future phase.
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