Overview The aim of our project is to develop a computational tool solving pathway engineering problems with strong practicability and great interactivity. In order to achieve this goal, we did a great deal of human practices with researchers, iGEM teams and social communities throughout our project. Silver "The most critical part of designing any solution is to understand the problem you're trying to solve." To become that intimate with the problem, we devoted ourselves to interactive with our software’s end users -- the very people who have this problem -- to understand their workflow and how they think our solution should work. Moreover, we communicated with peers who have the experiences of developing software. From these communications, we got a great deal of suggestions about how to build a software that is user friendly, beautiful, responsive, reliable, secure, and scalable. Interview with Prof.Liu Qi At the early stage of our project, we had an interview with Prof. Liu Qi to learn about how bioinformatics could benefit the research of biology field. Prof. Liu Qi currently conducts bioinformatics research at the School of Life Science and Technology of Tongji University. His biomedical Big Data Mining team (BM2, stand for Biological and Medical Big Data Mining) is dedicated to the application of evolving artificial intelligence and machine learning technologies to mine high-throughput biometric data and drug data for biological problem mechanisms. Fig. 1. Interview Prof.Liu and learn about applications of bioinformatics. During one hour communication with Prof. Liu, we learnt a lot about the applications and approaches of bioinformatics in many aspects. Here is the general information we got. Fig. 2. Applications and approaches of bioinformatics in many aspects. Prof. Liu inspired us that development of bioinformatics software should encompass biological information acquisition, processing, storage, distribution, analysis, and interpretation that combines the tools and techniques of mathematics, computer science, and biology with the aim of understanding the biological significance of a variety of data. Biology Labs Visiting We visited some of the biology labs in School of Life Science and Technology of Tongji University to study general needs of researchers and knowledge of biosafety in wet lab. We found that most researchers view biological data from previous studies as a treasure. Utilization data rationally not only help experiment design a lot but also benefit discovery of biological mechanisms. This inspires us to develop a tool that gather, integrate and analyze data on a specific aspect in synthetic biology. Fig. 3. Interview with Assistant Prof.Zhang and learn about biosafety in wet lab. What’s more, we learned basic knowledge of biosafety in synthetic biology during these visits. It is important to use biosafety mechanisms such as suicide genes and nutrient dependencies to ensure the organisms cannot survive outside of the lab setting in which they were originally created. Also, conduction of regular reviews of the biosafety in laboratory settings, as well as strict guidelines to follow are indispensable. Interview with Prof.LI Ping in Tongji University We had an interview with Prof. Li Ping and a discussion with doctoral students in Prof.Li’s laboratory in May at the beginning development of our project.Prof. LI now conducts scientific research in School of Life Science and Technology of Tongji University. Her research group mainly engaged in the fields of microbial fermentation and synthesis of natural active substances which are very relative to pathway engineering. Fig. 4. Discussing pathway engineering with Prof.Li.She told us that there are many challenges when conducting a pathway engineering project. One major challenge is in the design of the pathway. She told us that there are many challenges when conducting a pathway engineering project. One major challenge is in the design of the pathway, which may require sorting through thousands of possible enzymes and reactions wherein all the parameters have different substrate preferences and kinetic features. It is not easy for researchers to search information efficiently due to a considerable amount of data available on the Internet. “It will save much time for us if you could provide a computational tool for designing and constructing pathway.” She said. Moreover, we had a discussion with doctoral students about the optimization of the heterologous metabolic pathway. They told us that the most important factor should be considered is the metabolic burden in the pursuit to identify an optimal metabolic pathway. “Minimizing the number of heterologous reactions that need to be added would help reduce the metabolic burden.” They said. After an in-depth discussion with Prof.Li and her doctoral students, we learned that researchers engaged in metabolic engineering have a great demand for a computational tool that help them design and construct pathways. Moreover, they inspire us to consider more about metabolic burden when ranking pathways. iGEM Eurasian meet-up It’s a great pleasure to participate in iGEM Eurasian meetup 2018 in July this summer. Through engagement in the interactive presentation and excellent workshops, we learnt much about biosafety and bridging bioethics standards between continents. Besides these, we communicated a lot with other iGEM teams during the poster exhibition, which made us aware that there are a lot to be improved in our project to meet synthetic biology researcher's needs. More details can be found on our Integrated HP& Goldpage. Fig. 5. Our first day in Eurasia Meetup. Fig. 6. Many teams came and visited our poster. We exchanged a lot of valuable suggestions. Conference of China iGEMer Community, CCiC We attended 5th CCiC which was held by ShanghaiTech University in August this summer. During this meeting, we introduced our project to others through presentation and poster exhibition and gained lots of valuable feedbacks. Fig. 7. CCiC in Shanghai. Fig. 8. Our poster session in CCiC. Fig. 9. We gave presentation to other teams.Q & A section was really challenging for us. Here are suggestions from others. More details about how we improved our project based on these advices can be found on our Integrated HP& Goldpage. “In our simulation of metabolism with and without phase separation, we inevitably need the kinetic parameters of enzymes used in our pathway to produce β-carotene. So if your software could offer the accurate enzyme kinetic data, it will help us a lot make tenable predictions based on our models.” ——Peking “The illustration of algorithm can be further improved to make it more appealing; the items and parameters of the software can be explained more clearly for the audience to follow; a specific application scenario can be added to the presentation to make the whole story more complete and clear.” ——Some advices on presentation from judges Enterprises visiting To improve our project and make it serve the industrial community better, we visit three biotechnology and drug companies, Cathay Industrial Biotech, WuXi AppTech and MEIHUA Group. During these visits, our aims are as follows. 1.Studying the needs of industrial community 2.Seeking suggestions for improvement from the company's professional R&D staff 3.Promoting our web application to company staffs We had gained lots of valued suggestions from professional R&D staffs and improved our software based on the advice afterwards. More detailed information can be found on the Integrated HP& Gold page. Moreover, we generalize our tool and some of the R&D staffs planned to use it in industrial small molecular compound production design, which encouraged us a lot. No matter how it works well, it will be futile if no one use it. So promotion is important. Integrated HP & Gold An excellent project is inseparable from careful and creative thinking about whether your work is responsible and good for the world. To achieve this goal, we actively engaged with stakeholders and professionals to get the best out of our project. We integrated the extensive feedback we received into the design and evolution of our project. This page describes how the investigation of our HP issues has been integrated into the purpose, design, and execution of our project in details. It consists of two parts, Functions Integration and Project Assessment. Fig.10. Overview of Integrated HP Functions Integration Advancements in metabolic engineering have enabled us to engineer and express enzymes and construct novel pathways for various applications including drug discovery and value-added biochemical production. However, it’s quite a challenge to design and reconstruct a pathway with high yield and productivity in certain organism. So the very initial idea of us is to develop a computational tool for pathway recommendation. However, it is not clear that how to evaluate the pathway, what the general pathway engineering workflow of experimenters and how they think our solution should work. 1.Database integration: Inspirations from Prof. Liu and Peking: Prof. Liu emphasized the comprehensiveness and reliability of data used for pathway searching. He made us aware the importance of data. We obtained data from reliable database online, cleared redundant data afterward and integrated them to our software. “Data is the foundation of your software. Only by ensuring their accuracy can your software work well.” Prof. Liu said. We met Peking on the 5th CCiC meetup. Since they need the kinetic parameters of enzymes used in their pathway to produce β-carotene, it inspired us we can offer comprehensive information of compounds, enzymes, reactions and organisms to users to help them design experiments better. Data integration: Fig.11. Databases we integrated in Alpha Ant. Information display in Alpha Ant Fig.12. Result page of Alpha Ant. 2. Pathway Ranking Criteria Integration Inspirations from Key Synthetic Biology Laboratory of Chinese Academy of Sciences visit: Thanks for Prof. Li Ping’s referral and we got the opportunity to visit Key Synthetic Biology Laboratory of Chinese Academy of Sciences. We introduced our project and they gave us a valued suggestion on pathway ranking criteria from the experimenter's point of view. Researcher Yang Sheng told us generally the designed pathway for value-added biochemical production should coincide with the central metabolic pathway of chassis cells as much as possible to get high yield according to the experience in experimenting. It inspired us that we could design a scoring method based on the coincidence of the pathway to be evaluated and the core metabolic pathway in chassis cell. Fig. 13. Introduce Alpha Ant to Prof.Yang and Prof.Li in Key Synthetic Biology Laboratory of Chinese Academy of Sciences. Fig. 14. Prof.Yang show us around and help us get to know more about metabolic engineering. Improvement in pathway ranking: We integrated the new pathway ranking method into our previous criteria, which meets our users’ needs better. Also, we found that the verified pathways in papers had a higher score in software assessment.Check it in Validation. 3. Organism recommendation Inspiration from iGEMers: We attended the iGEM Eurasian meet-up in August and communicated a lot with other iGEM teams. Through those communications, we learned that most of them faced the problem of organism selection in pathway engineering. It inspired us to add a new function, organism recommendation to make our software more practical. Fig. 15. Other iGEM teams came to our poster and proposed advice to us. Improvement: Based on this human practice, we develop a model to scoring each microorganism. After ranking all those score, we provide users with top five organisms to choose. At the same time, related information about organism and pathway are optional to get. 4. SMILES Comparison Inspiration from R&D staffs in WuXi AppTech: Original thinking about SMILES comparison is derived from our visits to WuXi AppTec. Experts of WuXi AppTec proposed an idea to us. They said that sometimes their company got or designed a novel compound which did not exist in current database, and they want to find a possible way to synthesize it. So it came to our mind that what if we could compare the similarity between different compounds and select the most similar compound as a trigger to help us design new synthetic pathway, which can be very useful in small molecular drug discovery and synthesis. Fig. 16. Original thinking about SMILES comparison is derived from our visits to WuXi AppTec. Experts of WuXi AppTec proposed an idea to us. Improvement: We added a new function called SMILES Comparison into Alpha Ant. First we convert user’s input SMILES into molecular fingerprints by using RDkit toolbox. Then we compute similarity score between input compound and compound in databases by comparing their fingerprints. At last, we offer the similarity score and ranking list to users. Fig. 17. SMILES comparison page of Alpha Ant. Software Assessment: Results validation： We visit Shanghai R&D center of Meihua Group in September. Meihua Group is one of the largest MSG manufacturers in the world. It is the largest amino acid producer in China and an important member of the global bio-fermentation field. Their R&D center is dedicated to do research in amino acid production by bio-fermentation. There are several mature production lines for amino acid, which utilizes pathway engineering. We used these efficient pathways to evaluate the results of our software. Fig. 18. We visit Shanghai R&D center of Meihua Group in September. Meihua Group is one of the largest MSG manufacturers in the world. Aspects to be improved We visit R&D center of Cathay Industrial Biotech in September. Cathay Industrial Biotech focus on the industrialization of bio-manufacturing technology and practices, which have already achieved large-scale industrialization including long-chain dibasic acids, biobutanol, biopentylamine, and bio-based polyamides. Fig. 19. We visit R&D center of Cathay Industrial Biotech in September. We had an in-depth discussion with R&D staffs. They pointed out many aspects in our software to be improved based on their years of research and development experience. Due to lack of time, we decided to made future plans as follows. Future plan1: The optimum conditions for each reactions’ enzymes may be different, we will take into accont the positions and optimum conditions of each reaction’s enzyme when ranking pathway to guarantee that the recommended optimal pathway will achieve the highest efficiency under the same condition. Future plan2:There are other factors should be taken into account when evaluating the pathway, such as primary metabolite or secondary metabolite,growth phase or fermentation period of microorganism. We learned that in the production of industrial compounds, producers tend to use microbial fermentation in an anaerobic environment to reduce the biomass energy consumption and achieve high productity. So we will take the types of metabolite and growth state of microorganism into account when optimising pathway. Future plan3: We will consider about modyfing existing enzymes to improve efficiency of pathway. Enzymes are often engineered to carry out new functions in the context of synthetic metabolic pathways. Enzyme modification is a challenging and promising solution for improving pathway efficiency. We are planning to read related literature and develop this excitiing funcion afterwards. Education & Public Engagement Game: "GO ANTS!!!" Be Curious about the fantastic biochemical reactions in organisms? Feeling boring in biochemical textbooks? If your answer is yes, then try our game! This year our team designed and developed a fascinating game called “GO ANTS”. This game aims at spreading basic knowledge of metabolic pathway in organisms to general public. It is a double-operated game which incites people’s pleasure of cooperation.The players need to convert a compound to another compound through several enzyme reactions within a specific time limit in order to win. Players control two ants to move and collide with specific enzyme to complete an enzyme reaction via printing different keyboards. If the ants collide with wrong enzyme, then they need to collide with a specific enzyme to undo this reaction. People will learn related knowledge about enzyme reactions and metabolic pathways in organisms.A number of students enjoyed playing our game. Some of the players also gave some advice to improve the game experience. "This game is a litter difficult to play but with great fun!" by Sun "It makes the biochemical reaction no longer boring!" by Gao More excited it will be if add some inspiring music!"by Yi by Yi Fig. 20. Our teammate Xiaotian was introducing "Go Ant" to middle-grade students and they liked it! Lectures for Freshmen and High School Students Spreading a basic understanding of synthetic biology to the next generation is always what iGEM encourages us to do. So we held several mini lectures for freshmen and high school students this year. We introduced basic knowledge of synthetic biology and our project Alpha Ant. These lectures incited their interest in synthetic biology a lot and they also shared their creative ideas and interest of synthetic biology at the end of the lectures. Fig. 21. We held leactures in No.1 High School Affiliated to Tongji University. Fig. 22. Promoting synthetic biology to freshmen in Tongji University. Experience Sharing Session All of our team members have gained great self-improvement during the whole project development. We had the desire to share knowledge and experiences we have got to others. So we held an experience sharing session with students majored in bioinformatics. We introduced our project to them and shared our stories about difficulties we met and how we got over it. Fig. 23. we held an experience sharing session with students majored in bioinformatics. Several of the listeners who now conduct innovation and entrepreneurship project said they got many creative ideas about how to develop bioinformatics tools that benefit the biologic research. Moreover, they expressed their suggestions on our project, which also benefit our project a lot.