1. Competition Deliverables
c)Project Promotion Video
All required materials were submitted before deadlines.
We wouldn’t have accomplished our works without our team members’ efforts and assistance of other people! Special thanks to individuals and groups that have helped us are expressed on Attributions page.
3. Project Description
Acne, a common skin disease generally related to pores troubled by excess sebum blocking and Propionibacterium acnes proliferation, almost bothers everyone but has no particularly effective treatment. To address this long-standing challenge, our project aims to develop a novel acne therapy. We engineered a safe, nutritionally deficient Escherichia coli strain, which could degrade fatty acids in high efficiency and secrete strain-specific bacteriocin, for ameliorating sebum blocking and inhibiting P. acnes growth, respectively. In addition, a fatty acids-sensing system was constructed to enable precise therapeutics actuation only in appropriate conditions, minimizing the side effects of gene leakage expression. Furthermore, a high-throughput microfluidic chip was manufactured for accelerating the experimental cycle of directed evolution. It is anticipated that the Acneraser will build new bridges between acne research and synthetic biology, letting this promising and exciting scientific field change our lives from the very tiny things around us all.
We have added new information learned from literature and the experimental characterization to the previous parts (BBa_K3040118 and BBa_K861050). Furthermore, we have documented the method to culture Propionibacterium acnes with the help of China Center for Type Culture Collection（CCTCC）. In addition, we manufactured a high-throughput microfluidic chip for directed evolution and documented the 3D printing process of the hardware, assisting the construction of this microfluidics chip. Some Human practice, Model and Entrepreneurship work are done and we hope they will inspire the following teams.
All of these may be helpful, and we hope it will make some contribution to the iGEM community. Detailed contributions are available on Contribution page.
1. Engineering Success
We have accomplished Design-Build-Test-Learn (DBTL) cycles in Inhibition Module, Fatty Acid Sensing Module, Fatty Acid Consumption Module and Safety Module.
In the Inhibiting Module, we designed a part (BBa_K3763023) using the original Sec signal peptide to attain the secretion of PctA. When the feedbacks revealed the incompatibility with our chassis, we changed it into OmpA and created a new part (BBa_K3763024) along with future experiment plan to test its ability.
In the Fatty Acid Sensing Module, we used pFadD_Lac to make our engineered bacteria sensitive to fatty acid. We failed many times in inducting GFP but finally made it and explored the suitable way of testing the sensing efficiency, which was hoped to work in further study.
In the Fatty Acid Consumption Module, decompose fatty acids quickly is our goal. Only it didn’t work effectively as we over expressed the oxidases, when we found the incorrect enzyme expression. Then we changed to another strain and got positive results.
In the Safety Module, we struggled to construct amino acid deficient Escherichia coli. by knocking out hisBCD or glyA gene. Without positive results, we still starved in many ways and asking for PIs, they provided constructive ideas and expected to fulfill.
We have characterized several new basic parts in engineering this year: BBa_K3763021 and BBa_K3763042. We have also designed and built several composite parts. All of them are tested and clearly documented on the Registry. For three core basic parts:
BBa_K3763021 is the coding sequence of PctA, a bacteriocin aims at Propionibacterium species, including Propionibacterium acnes, the pathogen of acne. We’ve successfully expressed and purified precursor PctA with this part, and even though we haven’t prove its antibiotic activity, using PctA to inhibit s is of great potence and has its own advantages.
BBa_K3763042 is a coding sequence of fatty acyl-CoA dehydrogenase (FadE), a β-oxidation related enzyme. We overexpressed it in engineered bacteria and collected some data on the fatty acid decomposition ability of these engineered bacteria.
We founded 1 alliance—Skin Alliance and participated in 3 iGEMer alliances — the CCiC, the ICII and the Huazhong Alliance, in which we worked as a leading factor throughout in this season. We have met with a lot of teams, schools and organizations, established partnership and worked as groups. We held meetings, published booklets, did works of popular science and exchanged ideas on design, modeling and lab work. All the Collaborations are shown here.
BBa_K3763023 is a composite part to express our interested protein Propionicin T1 with native peptide and 6xHis-tag in cytosol, which can be purified using Ni-IMAC.After purification to test the inhibition ability of PctA against Propionibacterium acnes.
3. Human Practices
Integrated Human Practices is an overview of our human practices work which includes, Interview, Education and Inclusivity, we also made tight connection with some innovative teams and established some meaningful alliances.
In Integrated Human Practices, we first arouse this idea by word cloud and then gathered a large spectrum of suggestions from different professors, doctors and other skateholders to evaluate and reform our project, which made our work precisely targeted at the problem of acne; In Education, we published popular science booklets as e-books and paper books, and we delivered lectures about synthetic biology, skin-keeping and acnes for people of all ages, which aimed to promote mutual learning and discussion; in Inclusivity, we gained and analyzed pubic concerns about multiple aspects, including the treatment, concerns, side-effects, probiotics, products, synthetic biology and our project.
4. Proposed Implementation
The proposed users of our active dry probiotics are patients troubled by acne, especially those young people that often have a longer sick time. As a product applied on patients, the safety is a main concern and we have taken it into account in terms of lab work, project design and transfer. To implement our project to real world, multiple researches and interviews are done and we have designed a complete commercialization pipeline. You can get more information about the application attempt on Implementation and Entrepreneurship.
1. Integrated Human Practices
There are 5 parts in our integrated Human Practice. And Human Practice is an overview of our work.
In Interview, we visited experts and other stakeholders of various fields and adjusted our project with their suggestions, which covered a large spectrum from cell-free techniques to entrepreneurship experiences;
In Education, we set a gradient set of science popularization to people of all ages;
In Inclusivity, we stuck at market research and product transformation for better extension;
2. Improvement of an Existing Part
After comparing some existing RBS, according to the fact that most RBS with higher binding efficiency had the repetitive sequence of "AGGG", or "AAA" after the start base, we improved a new part of RBS(BBa_K3763000) and tried to compare it with the original RBS (BBa_B0030) to see which is better in some specific occasions. The performance of our fatty acids-sensitive module is improved to a certain extent.
3. Project Modeling
Our modeling part includes two subparts to explain how our engineering bacteria interact with P. acnes and how we accelerate fatty acid consumption of engineering bacteria, including growth simulator model and flux accelerator model.
Due to excellent cooperation, the model and the wet lab’s work are closely connected. The growth simulator model partially complements the experiment that we have difficulty in completing, and explains the feasibility of our project from an overall perspective. Additionally, results of the flux accelerator model has shaped our design in choosing enzymes. These are the details of our Model.
4.Proof of Concept
Our wet lab, hardware and model are combined together firmly to prove the feasibility of our design.
In the work of wet lab, we constructed a few parts to accomplish different aspects of our design, carried out experiments and proved the function of several of them.
In the hardware, we designed and constructed a hardware specialized for directed evolution.
In the work of mathematical model, we used mathematical modules to select the enzyme we need, and simulate the co-culture of our engineered bacteria and P. acnes, considering our project as a whole.
Detailed Proof-of-Concept is available here.
In this part, we met many like-minded friends, especially HUST2-China, we had similar target to solve the same problem in different ways, we have communicated with each other at least once a month, discussing about wet lab, HP, models and so on. The distance between us have made great convenience to do both online and offline work. The shining part is that we could give full play from our own school, such as there is a CCTCC (which was mentioned above) in WHU, we sent them some P. acnes to fulfill their wok. Detailed Partnership is available here.
6. Education & Communication
We have carried out different education activities aimed at people of all ages, including kids, senior school students, university students and the public. Education materials including a small game, popular science brochure and stickers are designed. In addition, based on Wechat, the public account allows comprehensive communication and interaction of articles, pictures, videos between our followers and us. Memes created by us will further enhance our influence.
You can read more information on Communication.
7. Excellence in Another Area
To overcome the difficulty in applying our iGEM project to real world, we develop an Entrepreneurship plan preliminarily.
In addition, we have made a pamphlet to popularize synthetic biology and skin related knowledge, a small game to better popularize knowledge to elementary school students, beautiful stickers, posters and memes to promote our project. Detailed Excellence in Another Area is available here.