I. Competition Deliverables
- Project Promotion Video
- Team Presentation
- Judging Form
II. Project Attributions
- Project Promotion Video
- Team Presentation
- Judging Form
II. Project Attributions
All of our team members have pulled together to complete this project for over a year. Wen-Liang Chen and Hsiao-Ching Lee served as our project instructors. We have received their guidance to the project in all aspects throughout the year. Jay Huang, Chun-Yuan Yun, Heng-Chien Liu, Yu-Tung Chen, Yun-Hsuan Hsiao, Ching-Hung Cheng and Zi-Yuan Lin were members of Wet Lab Group. They were in charge of papers research on Ubx proteins, codon optimization and gene design, experiment cycle design and conducting. Hsi-Jui Chang, Bo-Yi Jiang, En-Fu Liao, Kevin Liu and Shin-Hsuen Lin were the members of Dry Lab Group. They were in charge of predictive modeling, including protein expression model, anchor model, and color mixing models. Yi Tang, Li-Wen Wang and Wei-Hsin Chen were members of Human Practice. They were in charge of education, building science communication and a series of BioArt events.
III. Project Description
The initiative of our idea is to develop a versatile biomaterial with a wide range of applications. Ultrabithorax (Ubx), a transcription factor from Drosophila melanogaster we discovered, has become a great candidate to build the scaffold of the biomaterial. Ubx is biocompatible and biodegradable. It is able to self-assemble into nanofiber at the air-water interface by dityrosine bonds crosslinking. Furthermore, it retains its properties after fusing with other functional proteins. However, we found that current yielding of Ubx has not achieved satisfactory results due to stress response. Thus, we identified dityrosine containing motifs, functional domains Y167 and Y240 that contribute to the linkings forming self-assembly, and further engineered them with mRFP. The success of the functional test has further concluded that Ubx has the potential to successfully incorporate diverse functional proteins for wide applications in various fields.
To establish our color mixing model, we measured the fluorescent intensity to time relationship of Part: BBa_J04450 (RFP) and Part: BBa_K741002(gfp). With these parameters, we created a model for future teams to deduce the ratio of red and green for their desired color. This simulation shows that different RGB values lead to different color results. What's more important is that the changing degree of red and green values is based on our protein expression experiments. Thus, according to their desired color, future teams are able to speculate the specific incubation time for E.coli transformed with RFP and GFP genes.
On the contrary, our color mixing model also provides future teams a way to predict the color mixed by two fluorescent proteins. As future teams input the given RGB values and the opacity of different fluorescent proteins, our model enables them to visualize the mixing result.
I. Engineering Success
We conducted 3 engineering cycles to develop our novel Ubx biomaterial. In cycle 1, we created a composite Ubx part Bba_K4377006. SDS Page result showed the production was below expectation. In cycle 2, we developed approaches to narrow down to dityrosine containing domains among Ubx sequences, as dityrosine cross-linkings are evidenced to drive the assembly of proteins and peptides. ANCHOR algorithm and a model was used to precisely predict exact formation of dityrosine cross linking among the possible binding sites. Y167 region and Y240 of the Ubx showed the best probability of forming dityrosine cross linking. Thus, in cycle 3, we further engineered Y167 and Y240 with functional protein mRFP (Part:E1010), Bba_K4377007 in the attempt to verify the self-assembly function. Through functional tests, we had successfully achieved our goal of creating the biomaterial that contains self-assembling ability and preserving the function of functional protein fusion with them.
Especially forming a long-term partnership with NTHU_Taiwan. In our science communication project, we worked with KCL and NTHU_Taiwan to launch a series of bioart events, meanwhile, inviting KCIS_Xiugang and GEMs_Taiwan to hold in-person workshops in Taiwan. We also invited many teams to be our bioartists in our online exhibition. Aside from our initiatives, we were invited to participate in team CCU's cookbook writing for vascular disease prevention, team Tec Chihuahua's comic book drawing for young-kid education, team Ku Leuven's scientific podcasts recording, team GEMs_Taiwan's infographic translation project, team NIS_BIL_iGEM's video shooting to raise awareness about plastic pollution, team CSMU_Taiwan's iGEM theme song recording, and team KCL's podcast recording.
III. Human Practices
From forming the initial idea followed by executing our project plans, we always shaped our directions through human practice to ensure we’re on the right path. Initially, we have consulted academic experts specializing in material engineering who provided deeper insight to current biomaterial development. Aslo, we interviewed doctors with expertise in stem cell differentiation who informed us of potential applications in the field of biomedicine. Aside from academic guidance, we visited a fiber company and received some technical advice from the front-line manufacturer in industry. By receiving all the personal guidance from the stakeholders, we had developed a more comprehensive project.
IV. Proposed Implementation
In order to tailor our project design to the end users, we identified scaffolds for tissue engineering and regenerative medicine as our Ubx biomaterial implements. We decided to bioprint Ubx material attributing to advantages of 3D printing method - personalized design. Thus, we decided to conduct a measurement test of shear thinning to ensure the printability of our Ubx material. Aside from our image implents, we also envision others reengineering and redesigning our Ubx biomaterial for other applications other than scaffold. To visualize the idea, we developed the color-mixing model that can be adapted to simulate concentration of different proteins on the material. By the combination of our experiment and model, we are able to implement Ubx biomaterial in real life.
I. Integrated Human Practices
Academic and industry guidance allow for our full consideration of technical and ethical decisions required for the comprehension of the project. First, we consulted professor Ming-Jia Lee of Material Science for assistance on our structure design and fiber spinning. He suggested that we could alter the entropy of the protein and further launched a 3D-printing workshop opened to our team. Afterwards, we participated in his lab and learned fiber electrospinning and 3D-printer operation. Second, we approached doctor Huai-En Lu of Stem Cell Regeneration for biomedicine implementation. Guided by his advice, our drylab group developed a color mixing model to predict the effect regarding different protein ratios forming the biomaterial, which could be further applied in building scaffolds for cell generation. Lastly, we interviewed the CEO of Hung Chou Fiber Industry Co, Ltd. and he explained the production line of protein-based material in the textile industry.
II. Project Modeling
This year, the aim of our project is to create a novel biomaterial by engineering Ultrabithorax protein. We utilized models in our engineering cycles and further applied them to future implementation. Prior to engineering cycle 1, we used protein structure prediction model to predict Ubx protein structure to obtain better insights associated with Ubx protein properties. Yet, the protein expression of the Ubx is below expectation. In engineering cycle 2, we further narrowed down to dityrosine containing domains among Ubx sequences, as dityrosine cross-linkings are evidenced to drive the assembly of proteins and peptides. Thus binding region prediction model was built to precisely predict the formation of dityrosine cross linking among Ubx sequences. In engineering cycle 3, in the functional test we developed a tyrosine binding model to predict the best circumstance for the formation of dityrosine bonds. Eventually, we built a color mixing model for our future implementation in bioprinting a scaffold for tissue engineering by analyzing the relationship between protein concentrations and the output color.
III. Proof of Concept
In order to tailor our project design to the end users, we identified scaffolds for tissue engineering and regenerative medicine as our Ubx biomaterial implements. We decided to bioprint Ubx material attributing to advantages of 3D printing method - personalized design.
In professor Ming-Chia, Li's lab, we did measurement of viscosity for our protein sample and confirmed its shear thinning behavior. This proves that our UBX biomaterial has great potential as 3D printing material.
We further bioprinted our Ubx biomaterial, and had successfully printed out a plane layer. Below is our result.
We collaborated and formed a partnership with iGEM team NTHU_Taiwan in the project period. In march, both of our team members exchanged the initial ideas of the project , and we were inspired by the comments from each other. In April, our model group provided them with methods and useful software to model the degreagaion curve of the protein. Both of our team were joining AFOB 2022 ARS events in June. Days before the conference, we presented our slides to each other and made adjustments on the content based on the advice given. In July, NTHU_Taiwan invited us to record a joint episode in NTHU’s synbio podcast. In August, we helped NTHU_Taiwan conduct protein LL37 antimicrobial functional tests on E. coli. In September, We organized a BioArtist online public lecture event together. Furthermore, we invited NTHU_Taiwan to join our online BioArt exhibition to build science communication.
V. Education and Communication
Our science communication projects consist of the collaboration of interdisciplinary fields of art, technology and synthetic biology, providing interactive and engaging information to the public. First, we developed #BioArt_TheThirdSpace, working with KCL and NTHU_Taiwan to launch a series of BioArt events including BioArtist public lectures, BioArt workshops and an Online BioArt exhibition. Our digital BioArtwork created by the color mixing model Digital Fluorescent was exhibited in our BioArt gallery and in iGEM BioArt virtual gallery held by iGEM BioArt Steering Group. Second, in our #EdTech_GameSchooling we developed 2 virtual Role Play Games and 2 card board games to make difficult content more accessible to people without scientific background. Lastly, in our #SynBio_Lecture Material & Ed Events we established an education package containing lecture documents and slides. Those materials were delivered in our 3 education events held in highschools and were able to be downloaded online.