Development and research until March 2019
Since a few of our team members’ relatives enormously suffer from Type 2 Diabetes Mellitus, we, as a team, decided to focus our efforts on improving their lives. The most important issues that were brought up by the affected were severe side effects and the need of daily injections. As a result, we decided to look into the field of engineered probiotics and their possible application for the threatening issue of the world wide Diabetes epidemic. Consequently, after further research, we chose E. coli Nissle 1917 as our platform organism due to its unique characteristics and beneficial use as probiotic (Project/Nissle).
Furthermore, we decided to focus on early to intermediate stages of Type 2 Diabetes mellitus, hoping to lift the burden of daily injections and cumulative oral uptake of drugs (Project/Description). In this major group of Diabetes patients, most people are treated with Metformin, while new therapeutics like GLP-1 analogues continuously rise in their importance (Human Pracitices/Entrepreneurship).
Contacting Prof. Timo Müller
We contacted Prof. Timo Müller from the Helmholtz Institute in Munich (Human Practices/Experts), who helped us determine that Glucagon like peptide 1 (GLP-1) itself is not suitable as a therapeutic. Therefore, we researched GLP-1 analogs and finally decided on Exendin-4, a GLP-1 analogue originating from the lizard Heloderma suspectum. The human analogue to Exendin-4 is GLP-1, which is secreted from the L-cells in the small intestine when nutrients are available in the gut. To further mimic the natural state of incretin mimetics, we developed a glucose-sensing Catabolite repression - Tet-Repressor system (Parts/Parts Overview).
Additionally, Prof. Müller made us aware of the importance of bioavailability. We consequently researched secretion peptides and cell penetrating peptides to make sure our therapeutic would reach its target, the pancreas.
Meet-up with Prof. Andreas Fritsche
We met with Prof. Andreas Fritsche, deputy director of the Institute for Diabetes Research and Metabolic Disease of the Helmholtz Institute Munich at the Eberhard Karls University Tuebingen, to gather information about Diabetes patients and their willingness to use a genetically engineered organism as a therapeutic. In general, Prof. Fritsche explained that many people affected by Type 2 Diabetes especially struggle with obesity, which is often the cause for the condition but also comes along with the Insulin-treatment in late stages of Type 2 Diabetes. Thus, the idea of using GLP-1 analogues, which are known to support weight loss, was confirmed by this.
Moreover, Prof. Fritsche pointed out that, especially in Germany, the stigma concerning GMOs is quite large. Consequently, we decided that biosafety was the key to implementing living GMOs as therapeutics. Moreover, we were now convinced that human practice would be very important in our project, especially to address the negative perception of GMOs by the society (Human Practices/Overview, Human Practices/Education & Public Engagement).
Implementation of the CRISPR/Cas3 based kill switch system provided by Dr. Pengfei Xia
Integrating the public’s concerns about using GMOs, and to make our probiotic compatible with both the iGEM Biosafety standards and the foundation provided by German and European law (Safety,Entrepreneurship), we researched biocontainment. Finally, one of our supervisors, Pengfei Xia from the research group for Environmental Biotechnology supervised by Prof. Lars Angenent, helped us develop a CRISPR/Cas3 based kill switch, constraining the engineered probiotics to the assigned working place (Project/KillSwitch).
Researching permissible conditions for our Kill Switch
To determine the permissible conditions for our kill switch, research on bacterial regulatory systems was necessary. We used both publications and data previously generated by other iGEM Teams.
Soon, we realized that in 2017, Team AQA_Unesp had already tried to target Type 1 Diabetes Mellitus with a probiotic. Thus, we adapted several ideas from their project to implement the features of our kill switch (Project/KillSwitch).
Meeting with Synovo to talk about the business side and the project
To get further input on our project and first insights into a biotechnological company, we contacted Synovo and were happy to meet with them (Human Practices/Entrepreneurship). At Synovo, we got insight into the pharmacological side, as well as information about the implications of founding a start-up within the biotechnological sector.
Furthermore, we discussed the drug admission process up to clinical trials and a possible market-entry of our product (Human Practices/Entrepreneurship). Moreover, we talked about quality control and safety changes to the final product, for instance the removal of antibiotic resistances and the integration of the kill switch into the genome to make it more reliable.
Biosafety consulting with Dr. Jörg Schibel and Dr. Brigitte Walderich
Since we required CPPs (Project/Overview) for our project, we wanted to discuss potential safety concerns with safety officials of the University of Tübingen and the Max Planck Institute Tübingen. Together, we evaluated CPP safety, especially during our work in the lab (Safety, Human Practices/Experts) and decided to research and test diverse CPPs for our project (Project/Model).
Survey presentation at the Science Slam
To include the public in our project, we had a brief talk at a Science Slam hosted by the Medical Faculty of the University Tübingen (Human Practices/Overview). Here, we presented our project to a huge audience and kick-started our survey about the public’s knowledge on Diabetes, its treatment and the perception of GMOs (Human Practices/Survey).
The conclusions we drew from the survey were that not much is known about GMOs and Diabetes, thus we decided to host an information booth about Diabetes prevention and the potential of GMOs in the food and pharmaceutical industry later in the year (Human Practices/Education & Public Engagement).
From May 2019
Collaboration with the Institute of Innate Immunity Bonn (AG Eike Latz)
At the iGEM Spring Festival organised by iGEM Düsseldorf, we heard a talk about Diabetes and its prevention by the Director of the Institute of Innate Immunity Bonn, Prof. Eike Latz (Human Practices/Experts). Thus, we contacted him and asked for feedback and support for our project.
We established a cooperation for Diabetes prevention and were referred to his colleague Dr. Anette Christ. Anette was a great support, providing us with research data about chronic inflammation, GMOs and successful events for Diabetes prevention and health nutrition. Knowledge, which we used to design a flyer in collaboration with iGEM TU Dresden (People/Collaborations). This flyer, we planned to later hand out at our information booth and our panel discussion at the Experimenta in Heilbronn (Human Practices/Experimenta).
From May/June 2019
Intense WetLab work at the Wohlleben Labor
Our projects required intensive cloning of multiple regulatory elements (Project/Notebook). For the parts, we planned to use Biobricks that were sent to us with the 2019 shipping, Biobrick sequences from the iGEM’s database, as well as new sequences unique to our project. For the cloning, we used E. coli DH5alpha cells, as they allow for easier manipulation. As mentioned above, our final organism was E. coli Nissle 1917. However, we did not have a lot of knowledge about the probiotic and also found that, within the iGEM competition, E. coli Nissle 1917 was not properly characterized yet.
Modeling and characterization of E. coli Nissle 1917
To gain knowledge on our chassis organism, we decided to make use of a metabolic model of E. coli Nissle 1917. Due to the fact that no metabolic model of E. coli Nissle 1917 was available yet, we contacted Prof. Dr. Andreas Dräger who is an expert in metabolic modeling. Professor Dräger introduced CarveMe as a novel tool for the creation of metabolic models to us and gave us very valuable tipps for our subsequent modelling approaches. Inspired by our Advanced Transcriptomics lectures by Prof. Dr. Kay Nieselt, we decided to characterize the transcriptome of E. coli Nissle 1917 under various stress conditions. As a result, we required a vast amount of data generated by the WetLab such as growth curves under different conditions (Project/Nissle).
Collaboration with iGEM Costa Rica and iGEM TAS Taipei on the SDGs
The United Nations’ (UN) Sustainable Development Goals (SDGs) (Human Practices/SDGs) aim at improving the world’s situation by promoting prosperity, equality, peace and care for the environment. Altogether, the UN has set 17 of these goals, which need to be achieved by 2030. During a meet-up in Germany, a member of the iGEM Team Costa Rica approached us with their interest to promote the SDGs, as after all, iGEM projects generally aim to improve people's lives, while having great exposure to their communities.
When iGEM TAS_Taipei then launched their promotion for their video online conference about the SDGs, we decided to join forces and start a collaboration, called iGEMxSDGs (People/Collaborations). The challenge was launched in July 2019 and the proposal for an SDG integration as a bronze criteria to iGEM HQ was sent afterwards.
Meeting with Prof. Oliver Kohlbacher
To further refine our modelling approaches, we met with our long time supporter Prof. Dr. Oliver Kohlbacher of the Applied Bioinformatics Group at the University of Tübingen. During the meeting, the possibility of conducting a time-series RNA-Seq experiment to evaluate the efficacy of our kill-switch mechanism came up. Admittedly, we quickly discarded this idea due to a lack of time. Therefore, we discussed our RNA-Seq experimental design with respect to an E. coli Nissle characterization, evaluation of the safety of our tool as well as the viability of our regulation system under different conditions (Project/Nissle).
iGEM Amazonas Brasil (Manaus) Nissle Collaboration
Team iGEM Amazonas (Brasil, Manau) contacted us via social media, since they wanted to work with E. coli Nissle 1917 in their project but needed further knowledge on the bacterium. Thus, we organised a skype call and sent them our protocols, as well as the data we acquired during our Nissle characterisation (People/Collaborations).
Implementation of Collaboration with iGEM Tel Hai Migal on SDGs and ethical evaluation
iGEM Tel Hai Migal has announced a collaboration on the SDGs on the collaboration proposals of iGEM. We really liked their idea of an Escape box (People/Collaborations), since it is a really good example for the engagement of the SDGs within iGEM. Moreover, we discussed the ethics parts of their project (Parts/Download). To evaluate iGEM Tel Hai Migal’s project, we contacted Stefan Gammel (Human Practices/Experts).
Move to the ZMBP research group of Prof. Klaus Harter
Since we do not have a laboratory which we can use permanently, we moved one building across the street to the ZMBP. Here, we proceeded with our WetLab work and were happy to get to know a new surrounding and new groups which were happy to support our work.
Meeting with Dr. Rolf Hecker on Technology Transfer and IP
Since we were working on how an implementation of our project as a business would look like, we contacted Dr. Rolf Hecker (Human Practices/Experts) of the University of Tübingen’s Institute for Technology Transfer. Together, we talked about strategies for idea protection under iGEMs Creative Commons License and how a biotechnological start-up led by students could make it in the world of big pharma (Human Practices/Entrepreneurship).
Furthermore, we decided to form a stronger collaboration with the Institute of Technology Transfer, already talking about support for the iGEM Team Tübingen 2020.
Planning of RNA-Seq Experiments with Dr. Lisa Maier
September 2019 - Planning of RNA-Seq Experiments with Dr. Lisa Maier of the Interfaculty Institute of Microbiology and Infection Medicine (Human Practices/Experts), we designed our RNA-seq experiment and the conditions which we wanted to test (Project/Nissle). Moreover, Lisa kindly provided us with the anaerob mGAM media and bacterial supernatant of five species found in the human intestinal microbiome.
Anaerobe E. coli Nissle 1917 RNA preparation at the research group of Prof. Lars Angenent
For our RNA-Seq experiments, we required anaerobic bacterial growth and sample preparation. Thus, the research group of Environmental Biotechnology of Prof. Lars Angenent hosted our experiments and showed us how to work under anaerobic conditions. The prepared samples’ RNA was subsequently sequenced.
RNA-Seq with Angel Angelov of the NCCT
Due to the various conditions we wanted to test during our RNA-Seq experiments, we contacted Dr. Angel Angelov, one of the experts in RNA sequencing at the core facility in Tuebingen (Human Practices/Experts). With Angel and his colleagues, we isolated RNA, depleted rRNA and made RNA-Seq libraries which were afterwards sequenced.
RNA-Seq with EMBL
For the second half of our samples, the RNA Sequencing experiments were conducted at the Genomics Core Facility of the European Molecular Biology Laboratory (EMBL) in Heidelberg, under the supervision of Dr. Vladimir Benes’ group (Human Practices/Experts). For this, we previously had an initial meeting with Dr. Vladimir Benes at EMBL, discussing the planning of our experiment.
Moreover, at EMBL, we met with Dr. Daniel Machado, the developer behind the above mentioned CarveMe tool for metabolic model reconstruction to get tips on how to further improve our generated model. He further inspired our modeling by explaining and introducing Smetana, a tool which analyzes interactions in microbial communities. (Human Practices/Experts).
14th September 2019
SDG Video Conference organised by iGEM TAS Taipei
iGEM TAS Taipei from Taiwan organised an online video conference on the SDGs, in which we happily took part in since we already collaborated on the SDGs (People/Collaborations). For the conference, we made a video explaining how our team targets the SDGs (Human Practices/SDGs) and later discussed the SDGs with other iGEM teams participating in the conference. Overall, this highly supported the proposal of implementing the SDGs as a bronze criteria, since it showed that a lot of iGEM Teams are willing and happy to engage the SDGs within their project.
Student Lab/Panel Discussion Experimenta Heilbronn
The Experimenta is a science centre in Heilbronn, Germany, which aims at educating interested students from all over the country with exciting exhibitions and courses. Since we wanted to deepen the public’s knowledge about synthetic biology and advertise the natural sciences among young students, we collaborated with the Experimenta to supervise a student lab (Human Practices/Experimenta). Furthermore, in October, we organised and participated in a panel discussion about Biohacking at the Experimenta . Since the Experimenta is also interested in setting up a High school iGEM team themselves we will collaborate with them to start an iGEM team in next year's competition.
Microbiome experts talk
We were invited to present our project in front of the microbiome supergroup of the Max Planck Institute for Developmental Biology in Tuebingen. After giving the talk, we engaged in fruitful discussion and future perspectives of our project.
Testing iGEM Stuttgart’s new medium and using it for our characterisation
iGEM Stuttgart has developed a new bacterial medium, which we were happy to test in our Lab. Furthermore, we used the knowledge about the medium and the data acquired for our modeling.
Informative booth about the potential of GMOs in food and pharmaceutical industry
Together with iGEM Dresden and the Institute for Innate Immunity Bonn (Human Practices/Experts) we collaborated on pushing Diabetes prevention and advertising the potential of GMOs (People/Collaborations). Thus, on October 11th, we set up an information booth in the city centre of Tübingen, discussing GMOs and Diabetes prevention with the bypassers.
17th October 2019
Prize for exceptional student engangement
Each year, the University of Tübingen honours student groups for exceptional student engagement. This year, we were very proud to have won the prize for our human practice work. The award was presented to us during the University’s freshmen welcoming address, giving us the chance to introduce the students to our project, iGEM and the potential public engagement harbours for personal development.
28th October 2019
Since we received a lot of help during our iGEM experience - knowledge- and material-wise - we decided to say thank you to all of our supporters by presenting our final pitch for Boston to them. The pitch will take place on 28th October at the Max Planck Institute and will mark the finalisation of our project. Also, there will be cake.