Notebook
2021 September
Week 1:
Our iGEM journey began today. Our instructor Dr. Kuo Hui Ching gave us an introductory speech that highlights the potential impact of Synthetic biology. As the first iGEM team of Xiugang Campus of Kang Chiao, she encouraged us to aim for the gold and build up a solid foundation for the future iGEM team. She further delivered a clear vision and goal in the following year, indicating the overall timeline, the laboratory training, and the iGEM medals requirements.
Week 2:
For the second week of iGEM, we officially began the laboratory training. The first experiment we did was plasmid extraction. As a group of sophomore and juniors, today is the first time that we used the molecular lab equipment such as the micropipettes and centrifuge. Under the guidance of Dr. Kuo Hui Ching, we began to get familiar with handling the experiment by ourselves. It was a good start for the team.
Week 3:
Following what we did in the second week of iGEM, we continued the yeast culture protocol. We continued practicing inoculating yeast colonies and sterilized techniques. Dr. Kuo Hui Ching further guided us to conduct the protocol of stocking the yeast under the conduction of an 80-degree freezer.
Week 4:
In the last week of September, Dr. Kuo Hui Ching gave a lecture on highlighting the differences between the “in vivo '' and “in vitro'' projects. She also introduced a clear picture of cloning steps. For the laboratory training, we did the yeast genomic DNA extraction as well as the DNA agarose gel electrophoresis. This allowed the team to determine the plasmid concentration, checking the purity of the plasmid we got from the plasmid extraction we did in the previous weeks.
2021 October
Week 1:
During the first week of October, we continued our journey in learning the techniques in the field of Synthetic Biology. Our lecturer, Dr. Kuo Hui Ching, guided us through the principles and mechanisms behind bacterial transformation. We then got our hands dirty to perform the procedure on our own, and through that experience we learned about the importance of the step in cloning: without transforming the bacterial competent cells, DNA can not be cloned within the bacteria!
Week 2:
For our second week in October, we learned how to do bacteria plasmid extraction. It was in this week that we had our first experience with using phenol chloroform. Dr. Kuo was adamant about us having to wear all of the lab equipment when handling the chemical, as we later learned that skin exposure to phenol chloroform could cause itchiness and irritation. After learning of this unique interaction of the chemical with our bodies, all of us were extremely careful when handling it during experimentation.
Week 3:
After we learned about how to use restriction enzymes to cut the designated site for cloning, we learned that we need to digest these restriction enzymes to prevent it from interfering with further processes of cloning through the double digestion protocol (we used KPN1 and SMA1 as digestion enzymes). Another major part of the week was spent on troubleshooting since our previous bacteria transformation didn’t produce any colonies, we suspect that our cloning process wasn't successful or the competent cell used expired, so we plan on repeating the protocol again in the future but checking conditions of all materials more carefully.
Week 4:
During the last week of October we learned about the procedures of Polymerase Chain Reaction, or PCR for short. As students entrenched in the COVID pandemic, we were so used to hearing the phrase PCR, but we did not really understand how it works. Through listening to Ms. Kuo's lectures, we finally understood how it works and were surprised by how simple it was to understand the principles behind it. In addition to that, it was also surprising to see how the PCR plays a role in the cloning procedure.
2021 November
Week 1:
For our first week of November, we learned how to do plasmid transformation. During the procedure for plasmid transformation, Dr. Kuo explained to us why we needed to use both MgCl2 and CaCl2. Ms. Kuo gave a great analogy where she said that we wanted to beat the bacteria up so we can transform the plasmids, but not too fast so that it will die. That is why we had to use MgCl2 first to soften the yeast cells up, and then CaCl2 to help with the plasmid transformation.
Week 2:
For our second week in the lab, we learned how to run PCR products on DNA gel. While the majority of the class had heard and learned about DNA gel electrophoresis in biology class, setting up the actual kit and inserting the DNA was a first for many of us. The team in general struggled with the loading dye portion of the experiment, as we weren’t used to using pipetments to suck up solutions on kim wipes. But after repeating the process a few times, the team eventually got the hang of it.
Week 3:
In week 3, we learned how to inoculate single colonies of bacteria/ yeast onto a new plate. Everyone on the team was extremely excited to do this experiment as this was our first experience with the culture hood. Dr. Kuo explained to us in this class how the purpose of the culture hood was to create a sterile environment for cell culture experiments, and emphasized that by spraying our hands heavily with alcohol every time we were going to do an inoculation under the hood. Not to mention that everyone was extra careful when doing the experiments as Dr. Kuo stressed multiple times that the culture hood was one of the most expensive pieces of equipment in our lab and school.
Week 4:
This week was one of the few times that we saw Dr. Kuo was mad at us. The reason was that the whole class had forgotten to let the T4 DNA Ligase Buffer dethaw before adding it into the solution. When Dr. Kuo found out about our mistake, she was fuming, explaining to us how adding T4 DNA Ligase when it wasn't completely in liquid form would cause the solutions to differ in amount. She then told us to start the whole experiment over again, and we all made a mental note to let solutions dethaw before doing any experiments in the future.
2021 December
Week 1:
After learning the T4 ligation protocol, we learned the procedures to perform the bacterial transformation heat shock. Ms.Ching gave a lecture reviewing the protocols we learned. She highlights the techniques that help to determine the cloning works, ensuring that the target gene is cloned into the yeast.
Week 2:
In the second week of December, we conducted the experiments on isolating the colonies from pGal-GFP bacterial transformation plates onto a new LB-amp plate. We practiced how to inoculate the single colony on the LB-Amp plate by using the micropipette to determine the cloned pGal-GFP plasmid.
Week 3:
This week we practiced running the PCR samples on DNA agarose gel as well as setting up the PCR to double check insert DNA band. We also conducted the experiment of plasmid extraction from bacterial colony broths to send out sequencing. Our experiment result is shown on the diagram below. We found out that the long period of time of DNA inoculation resulted in the gel unable to express all bands of DNA, forming the degradation of the gene.
Week 4: Making yeast competent cells
For bacteria transformation, we had to learn how to make our own yeast competent cell after conducting cloning on yeast instead of bacteria. We also learned about the several benefits of using yeast for cloning compared to bacteria, including lower percentage of introns and sharing a lot of the same cellular process with the human, which makes it a better model for testing products for humans.
2022 January (Winter Break)
2021 February
Week 2:
Today, we have finally decided on our topic. At first, Ms. Parker and Ms. Ching divided all of us into four teams, in which each team was required to come up with a topic. After that, we will have a period to do research on the team topic and finish a presentation introducing our project. Then, once the team presented the projects, we will vote on our favorite topic. It turns out that obesity is going to be our objective for the 2022 iGEM Competition.
Week 3:
Based on the voting last week, we had decided our topic: obesity. This week, we will be focusing on finding the suppressor genes that will prohibit obesity, as well as the product to synthesize with the suppressor genes. In the end, we found out the two substances, SNF1 in yeast and AMPK in animal cells, that are related to regulating metabolism, a key factor of obesity diagnosis.
Week 4:
This week, we started to perform yeast transformation. Even though we have practiced our lab skills for a whole year, some of our members still make some mistakes, for example, protein contamination. However, as the experiment progressed, the team members started to feel comfortable with taking each step carefully and precisely.
2022 March
Week 1:
Following our work from last week, in the first week of March, we continued yeast transformation. Everything is finally getting on the right track, and the experiments are completed smoothly, despite some minor details omitted. We are progressing faster than we expected, which allows us to have longer preparation time for the competition.
Week 2:
Today, we moved on to the next protocol – cloning SNF1 into pGal Vector. Cloning is the most complicated protocol in synthetic biology, and many of us started to encounter difficulties. Ms. Ching had enough of our mistakes and demonstrated the experiment again. We were impressed by her skills and experience. After the demonstration, the members finally solved the problems.
Week 3:
After cloning SNF1 into the pGal Vector, we are now knocking the SNF1 out. This week, we decided not to make Ms. Ching angry again, so each of us worked to the best of our abilities. Through weeks of actually “experimenting,” we found ourselves picking up the pace of the experiments. Within one afternoon, we had our samples, which took us one entire day in the past.
Week 4:
This week, we did the most exciting part of the project: running the sample on gel. It is so satisfying to see the samples running through the electrophoresis gel. During our waiting time, we started to work on our wiki page. So exciting!
2022 April
Week 1:
At the start of April, we started the cloning process of SNF1 into pGal promotor. To achieve this, we started the process of cloning using restriction enzymes and PCR protocols we learned previously. This was the first time we started to conduct cloning independently and applying the knowledge we previously learned and integrating it into our current project.
Week 2:
After the cloning process, we ran gel electrophoresis to ensure the base pair number matches our design, and we sent the cloned product out for sequencing. The sequencing resulted with a 100% match with our vector design, so it is ready for bacterial transformation.
Week 3:
We conducted bacterial transformation this week after ensuring the cloning product was correct. By using ampicillin as a selection antibody on the agar gel plate, we ensured that the colonies grown all contained our cloning product. After seeing several colonies growing on the plate, we chose a few colonies and inoculated them for further experimentation.
Week 4:
We wanted to conduct RTq-PCR to observe if the SNF1 genes are really induced after cloning, so we had to extract RNA. It was the first time we did proper RNA extraction and faced some slight challenges in extracting the RNA instead of the protein parts due to our immature experimental techniques. But practice makes perfect, and after a few trials, we were able to successfully extract RNA ready for conducting RTq-PCR.
2022 May
Week 1& 2:
Some of the members didn’t come to school because of COVID-19 and AP Exams. The present members were focusing on wiki page preparation.
Week 3:
This week, instead of experimenting, Ms. Ching taught us the protocol for RNA extraction. Despite the complicated nature of the protocol, the team members were able to understand the overall process and purpose of the protocol. Furthermore, Ms. Ching talked about the concept of RT-qPCR, which took a huge part in our future experiments.
Week 4:
Today, we continued to delve deeper into the protocols and lectures. Ms. Ching introduced us to “truncated SNF1 cloning”.
Learn truncated SNF1 cloning and RT-qPCR technique
2022 June
Week 1:
This week we split into 3 groups to do the background research for the project. Group 1 focused on obesity, researching data about it. They further investigate the potential factors that caused obesity as well as its impact on human health. Group 2 focused on AMPKalpha research discovered in humans related to several stresses causing obesity and chronic disorder. They also research on the supplements that can boost metabolism to mitigate obesity in human society. They found green tea and brown algae. Group 3 focused on the benefit of using yeast to do experiments for human diseases.
Week 2:
In the second week of June, we completed the background research. The three groups shared their information to the class respectively. Ms.Ching also introduced the concept of Western blot. We watched the videos that demonstrate the procedures of Western blot.
Week 3:
In the third week of June, we learned the protocol to collect the RNA sample for RT-qPCR that helps us to ensure the pGal promoter is functioning properly. We also conducted the experiment of the yeast extraction. For the iGEM project, we began planning for the promotion video.
Week 4:
This is the last week of iGEM for this semester. We continued making the promotion video. We also separated the tasks that needed to be done throughout the summer vacation.
2022 July
Week 1:
This week, we began to collaborate with other schools: NYCU Formosa, Wego High school, TEC Chiwawa, Korean HS, Empire Gene. Online meetings were hosted to plan for future collaboration.
Week 2: 
Our arts team had produced two pages of comics introducing nobesity.
Week 3:
We worked on the 2022 KCIS x NYCU_Formosa Educational Workshop Collaboration Proposal. Due to the fact that many KCIS students are interested in joining the iGEM team, we decided to host a workshop introducing synthetic biology, our projects, and other team’s projects. We hope the workshop will give them a big picture about iGEM.
Week 4:
We finished editing the short video for NCHU to introduce our project – NOBESITY. Two of the best members of our team, Charlotte and Andrew, researched and edited the video all by themselves. Such an accomplishment.
2022 August
Week 1:
We continued collecting the data for our projects. We conducted the experiment of yeast RNA extraction and the RTq-PCR. We also began planning the information of the Wiki contents by organizing the information that we had collected throughout the year. The human practice team had a collaboration with the ASIJ iGEM team, planning an educational workshop that introduced synthetic biology and our projects to the dorm students in our school.
Week 2:
This week, we have met with the ASIJ_Tokyo high school. We were invited to an Taiwan Synthetic Biology Alliance conference, where we will have to do a presentation to share our project and exchange ideas with other teams. We are mainly preparing for the conference this week by making the posters and designing presentation layouts to make sure other teams can most effectively understand our project.
Week 3:
Last week, we finished filming our NOBESITY promotion video. This week, we have host a collaboration meeting with Wego highschool. Moreover, our promotion video team began to edit the video. Thanks to them, we now have an excellent promotion video. We watched the entire promotion video as a team and we shared each of our suggestions on areas of improvements and how to modify the content.
Week 4: 
This week, we selected four team representatives to attend the “Taiwan August Synthetic Biology Alliance Conference”. There, the group of four presented Nobesity and listened to project presentations of other teams in Taiwan. Moreover, we received feedback from and gave feedback to other groups. After the conference, we reviewed the feedback and began to improve our project according to it. We also started to explore possibilities in making our project into an industrial product to enhance the implications of our synthetic biology project in everyday life.
2022 September
Week 1&2:
This week we have a collaboration meeting with Korea_HS and Empire Gene. At 9/1 we co-hosted a bio workshop event with NYCU Formosa. The event was aiming to introduce Synthetic biology to ninth graders in our school. At the end of the bio workshop, we design a bioart experiment to give participants some idea about what synthetic biology is like. We use a more interesting way to present this idea, by drawing fluorescent bacteria (E coli. GMP) on agar plate. Through these experiments, participants can see the bacteria that are invisible to them in daily life.
For the human practice team, they had a collaboration meet with three schools: TEC Chiwawa, Korean HS, Empire Gene.
Week 3&4:
This week we have a meeting with NYCU Formosa about our collaboration. For the past two weeks, the team prepares the information required on the wiki page. Although a lot of information is still unfinished, our team sought out ways to maximize our productivity. In the end, we have finished half of the preparation.
2022 October
Week 1:
For the first week in October, we put our full strength to wrap up everything and complete the wiki website. The wiki website required tons of information, and the poor wiki team spent a lot of time helping us to update mistakes. However, thanks to the teamwork, it is coming together! In addition, we also conduct experiments of yeast survival assay to complete the final step to see if our idea behind – NOBESITY – is valid and reliable. We can only conduct it for 24 hours due to time constraints.
Week 2:
Most of the website is complete apart from a few edits, now the race has been on to complete our results page and proof of concept! The experiments team is going full steam and handing off pages and images to the wiki team to get all of it up in time. While we didn’t complete as many survival assays as we wanted, we did get a stress test that showed brown algae mitigates stress! After the final wiki uploads we will be on to completing out presentation and making sure the team has a good understanding of the project as a whole.
