Engineering as a problem-solving philosophy
Since the core of our team is composed of Biomedical Engineering Master’s students, we found it only natural to employ a problem-oriented approach throughout the project. For us, it was almost second nature to adopt the “engineering cycle” philosophy outside of the immediate dry and wet lab context. This ensured that the prototype design was not only guided by the dry and wet lab results, but also informed by the outcomes of the human practice initiatives, insights received from other iGEM teams, feedback from the more global scientific community, etc. Furthermore, the said outcomes of the lab work generated additional questions, which were quickly reflected by our integrated human practices, communications and changes to the overall strategy, as showcased by the implementation section.
Since almost the entire team was involved across multiple areas of the project, this integration was almost seamless, allowing us to work as a well-tuned mechanism - wittily represented by the rainbow gears (Although we could never agree whether it is pronounced .gif or .gif ...). To fully reflect our philosophy, we decided to split the mechanism into separate cycling gears, showcasing several examples of how the engineering mindset was generating ideas and insights both inside and outside our laboratory work.
iGEM’s introduction to the engineering process describes the design stage as the entry point of the engineering cycle. Yet, we felt like in the broader context of engineering as a problem-solving philosophy, it is crucial to start with learning the limitations of the existing solutions to the problem. For instance, previous teams have already designed several algorithms that generate toehold switch candidates for a target messenger RNA. However, these algorithms were not working for much shorter microRNA targets. Still, the access to their source code allowed us to better understand the approaches utilised for toehold design and quicker construct our updated model. Furthermore, depending on the packages and libraries employed, different code will produce different secondary structure and Gibbs free energy estimates. Both of these factors helped inform our first design, as we selected the candidates that performed well with different estimation methods. Finally, we tested our candidate libraries with external algorithms to double-check the result with an external model. This is where we discovered some inaccuracies in our code. For instance, our RNA energy estimates included the T7 transcriptional promoter sequence, which is obviously not right. We also had some trouble accounting for the reporter gene RNA impact on our genetic machine (a little diagram on the right is there to visualise the source of confusion), but eventually we managed to generate perfect candidate libraries for several target miRNAs. Once satisfied with the test, we could proceed to ordering the toehold sequences and preparing the wet lab experiments, which in turn informed the microfluidic design in the downstream dry lab efforts.
Of course, the dry lab cycle wouldn’t be as impactful in isolation. We chose two candidate libraries (toeholds targeting miRNA-141 and miRNA-21) and utilised two different methods to ligate the toehold sequences onto our plasmids. Our system design accounted for antibiotic selection, transfection markers, and selecting the bacteria capable of expressing genes under a T7 promoter. Unfortunately, neither of the ligation techniques worked properly, preventing us from generating sufficient data and achieving the PoC. Alas, as we were limited by the time remaining until the end of the competition, we could not continue the engineering cycle. Yet, we have designed the downstream steps, which involve a different selection of the restriction enzymes, a different bacterial strain, and using the candidate libraries redesigned with the improved dry lab algorithms. In the meantime, our experiments on exosome isolation from cancer cell culture and target miRNA quantification work have been more fruitful - feel free to learn more in the Results section!
Human Practices cycle
The key values of the Human Practices - Reflection, Responsibility and Responsiveness - line-up almost seamlessly with the iGEM engineering cycle. Reflection, the process of thinking through the team’s motivation and goals, is essential to the first steps of learning about the problem. Responsibility takes the spotlight whenever we design a new plasmid, build an educational workshop for a school, or consider the best implementations of our final product, as we aim to improve the lives of cancer patients around the world. We always ensure that the team stays faithful to the key iGEM values - respect, honesty, fairness, and others - as everyone works to minimise risks if having a negative impact. And, finally, we must be responsive to any problem arising from testing, and adapt the strategy based on the insights. For example, we spent a long time editing any communications aimed at the patients to remove potentially insensitive questions or messages. If you want to learn more about the impact of the human practice insights, and about the adaptations we have made throughout the project, please give the relevant sections a read - Human Practices and Implementation.
Social media, team meetups, science communication, educational work, and much more - communication behind an iGEM project is both extensive and diverse. Different key messages and target audiences require a tailored approach, which offers a large opportunity for practicing the engineering cycle approach. From the number of “likes” to the questions asked by potential sponsors, there is no shortage of feedback and insights to guide further design and building of the outreach strategies for different audiences. We learned how to adapt our presentations for different occasions, as well as what our social media followers want to see on our Instagram and Facebook pages. And, of course, the format and the content of our messages kept evolving throughout the iGEM journey. Don’t hesitate to check out our social media, education and communication sections to see how our presence has evolved both online and during in-person events!