Recent years have witnessed the CRISPR system enjoying greater and greater popularity. To address related technical problems and expand our way of thinking, a CRISPR-themed meetup was hosted by us and HainanU_China. On the meetup,7 other teams , including CPU_China, SCU-China, BIT DUT_China, HiZJU-China, NWU-CHINA-A, SCAU_China, and SJTU-software joined our extensive discussion. We also invited three special guests to give comments on our presentation and discussion. They are Yang Jianzhe, who acted as iGEM Ambassador in 2021, Liu Jingrong who has been tutoring more than 20 iGEM teams and ShiSonglin, who is a member of Biodiversity Leadership Program and familiar with CRISPR system. With 8 other iGEM teams and three special guests, we did extensive discussions on experiment, modeling, wiki, and human practice respectively.

For our part, after presenting our project in great detail, other teams gave us precious advice on algae cultivation, CRISPR application, eletro-transformation, and so on.

Minutes of our Q&A sessions
Q1: (Liu Zhirong)What is the schedule of your experiment?
A1: (UESTC_Biotech)The period of algae cultivation is long, but after exploration, we have been able to meet the requirements of synchronized and well-scheduled experiments . The experiment will not be completed until mid to late September.

Q2: (Liu Zhirong)What is the difference between oil production by microalgae and fungi and what are the advantages of using microalgae?
A2: (UESTC_Biotech) The purpose is similar between the two, but we have innovated the experimental route to find possible genes by using various stress conditions and then doing transcriptome analysis, followed by performing CRISPR knockdown to achieve the purpose of effective oil production.

Q3: (SJTU-Wei Jialu)Is there a prediction for the lipid output?
A3: (UESTC_Biotech) It is expected to increase 70-80% on the wide type of C.reinhardtti . Ideally, there could be 2-3 times more lipid output, and it depends on the amount of algae. To be specific,1g of original algae has 0.2-0.3g of oil, but we can upgrade it to 0.5-0.6g oil output.

Q4: (Luo Yuning from UESTC_Biotech) How to do lipid extraction?
A4: (SJTU-Wei Jialu) We had done lipid extraction in an innovative experiment by using the most traditional chloroform - dioxin (chemical reagent) extraction method , where the microalgae were grown normally in 1.5L shake flasks, but less than 1g of lipid was extracted from about 10 grams of algae. In general, oil production from microalgae is environmentally friendly, but the yield is low.

Q5: (Luo Yuning from UESTC_Biotech) Is there any component analysis of lipids done?
A5: (SJTU-Wei Jialu) No analysis was done because the extraction amount itself is too small to support the component analysis.

Q6: (Advisor, Shi Songlin) How is the increment of lipid output calculated, is there any evidence?
A6: (Luo Yuning from UESTC_Biotech) The results are what we got from the literature which uses gene silencing to achieve 70% to 2-3 times increment. And if we do combination knockout or gene knockout, we may get more amount.

Advice from special guest Shisonglin for our presentation

1. It is recommended to have references on the presentation. Not just images, but evidence or the process of inference.
2. The lipid extraction should be done as soon as possible to achieve the desired effect.
3. The purpose and content of the project are very significant for the world's energy optimization, but your team need to accelerate the experimental progress.
4. Gene editing technology could strengthen biosafety supervision.

After the meetup ended, a brochure for this meetup was formed to record the whole process of this meeting. We did extensive discussions on experiment, modeling, wiki, and human practice respectively. Through this CRISPR meetup, our knowledge of the CRISPR application has been broadened. Every team present had unique attainment on this meeting, and that is where the significance of communication and friendship lies.

Besides, we also attended CCiC(Conference for China iGEMer Community) to do the presentation of our project, as well as discussing widely with other 84 teams . On this conference, the judge said that our project has vital importance for current energy problems. But we should enhance biosafety levels on the original basis due to the application of gene-editing technology. Significant also, our team captain Wang Xingjie also acted as the judge to give comments and advice on other teams' projects.

Click the link below to learn more about DUT_China. After attending CCiC conference, we got to know that DUT_China also uses the CRISPR system. We exchanged the title and abstract of our projects through social media and surprisingly found that we were both doing the similar modeling for off-target prediction. We kept communicating during the process of constructing our model.

As our team uses CRISPR-Cas9 system to modify the metabolic pathway of microalgae, they gladly invited us to test our model for the prediction of off-target activity. Based on CNN, DenseNet, an outstanding neural networks model was used by DUT_China to screen out sgRNAs with high efficiency. UESTC_BioTech also developed a thermodynamics-based model to predict the potential off-target activity of the selected guide RNA.

The model built by DUT_China is very different from the model we constructed in terms of the overall objectives and the methods used, which leads to the fact that the results obtained by both parties cannot make direct comparison. Therefore, no quantitative analysis can be given. However, to a certain extent, what could be obtained is that both teams have similar results for off-target evaluation. Therefore, the correctness of our team's model was further verified by the model of DUT_China.

At the same time, during the communication between us, we found that our friends from DUT_China had some deviations from us in the understanding towards the concept of off-target. So, after further in-depth communication, we made our ideas and thoughts understood by our friends from DUT_China, which laid a good foundation for further collaboration. Then, we provided them with enlightenment on the algorithm design for finding potential off-target sequences in the whole genome sequence, and provided them with an optimized solution for the acceleration of their algorithm.

Click the link below to learn more about HainanU_China. In the middle of the project, we encountered HainanU_China. Based on the common ground of utilizing CRISPR system, we further communicated in terms of human practices, wiki building, and artwork layout, etc. We continued to improve our respective projects during these exchanges, and reached good partnership. It is definitely a wonderful and happy experience with HainanU_China.

During an online meeting, we learned that HainanU_China plans to use the CRISPR-13 system to construct of the nucleic acid detection platform, which is very significant in the global pandemic context and relevant to the technology used in our project. Based on the aspect of CRISPR technology, we reached a preliminary intention to deepen our communication.

We learned that many Chinese iGEM teams usthe e CRISPR system. In order to benefit wider community as well as discussing CRISPR related technology more comprehensively , we invited CPU_China, SCU-China, BIT DUT_China, HiZJU-China, NWU-CHINA-A, SCAU_China, and SJTU-software and other teams to share their projects. Besides, We also invited three speMid-termsts to give comment on our presentation and discussion. They are Yang Jianzhe who acted as iGEM Ambassador in 2021, Liu Jingrong who has been tutoring more than 20 iGEM teams and ShiSonglin who is a member of Biodiversity Leadership Program and familiar with CRISPR system. Apart from inviting teams, we often met online to settle down details of meetup. To be specific, we distributed tasks consisting of making invitation letters, designing application form, writing hosts word, noting down minutes and so on.

After doing full preparation beforehand, we hosted this meetup together on August 14. With 8 other iGEM teams and three special guests, we did extensive discussions on experiment, modeling, wiki, and human practice respectively and sought advice and solutions for one another, during which process we built closer friendships. These questions and answers, as well as the title and abstract of each team, were finally compiled into a brochure for future reviewing and experience accumulation of iGEM competition. The official account of CCiC community reposted our meetup summary to propagandize and recognize our efforts.

In the later stage of the project, we both encountered a lot of difficulties in terms of the presentation of the project to the public,the construction of the wiki, the layout of the artwork and the execution of Human Practice.The relevant persons in charge on both sides shared ideas and solutions, so as to achieve better results on respective projects. For instance, when HainanU_China got stuck in writing iGEM brochure for experience sharing, we offered them precious sample from our former iGEMer and joined them in the compilation of brochure for sharing iGEM related experience.

Click the link below to learn more about Sorbonne_U_Paris

During the early process of extensive communication, we are lucky enough to find our intimate partner Sorbonne_U_Paris.

We surprisingly found that we use the same chassis organism--Chlamydomonas reinhardtii. After meeting for the first time on Zoom to introduce our projects respectively, we learn about the project of each other and reached a consensus on establishing an in-depth partnership. With regards to the project of Sorbonne_U_Paris, their project this year is to use Chlamydomonas reinhardtii as a food source for treating iron-deficiency anemia.

Through extensive discussion on CCiC, our team decided to design a killswitch to enhance our safety level by designing a killswitch. But we lack experience in this aspect. After hearing our dilemma, Sorbonne_U_Paris warm-heartedly tutored the whole design of the switch.

While they were doing their project, they found it tricky to remove chlorophyll to make their algae tastier, under which circumstance we suggested the knockout of the PSY1 gene which is closely related to chlorophyll synthesis and provided them with its references, specific information, constructed plasmid and designed locus of sgRNA.

In the relative later period of our project, Sorbonne_U_Paris dedicated itself to constructing modelling for simulating the growth curve of C.reinhartii. And they needed data of the growth curve of C.reinhartii to complete their calibration. After learning that, we provided them with it to enrich their data for modeling construction.

To further deepen our collaboration and communication, we collectively did a research on the GMOs(Genetically Modified Organisms) regulations in respective country, and finally summarized the essence we learned in the form of an video. It is of vital importance because we can make our potential users of GMOs project learn about the policy context in different countries, which contributes to further education and communication in distinctive regions in the realm of GMOs.