1. Overview
An in-depth communication is enough for us to learn about other teams, however, we want assistance from close partners in the long term, which can continuously help us broaden our minds and perfect each other’s projects. Not only can we provide each other with suggestions on experiment and modeling, but also allow us to conduct social activity and ponder public feedback jointly for obtaining more valuable insights in human practice.
We have established a partnership with NUDT_CHINA since last year's competition, during which we have maintained close contact. Affected by the local epidemic, our early cooperation was mainly online, and it was not until September that we had the opportunity to hold two large-scale offline events.
2. Online meeting on 28th, July
We have had a online meeting on 28th July and discussed our projects and works.
As for our work in wet lab, The AIOD-CRISPR system binds two different sites by using a pair of Cas12a-crRNA complexes generated from two separate crRNA, close to the binding site of the primers in the target sequence. The AIOD-CRISPR system not only enables the detection of nucleic acids with few-copy sensitivity, but also addresses the limitations of the PAM sequence-dependent design of crRNA and expands the scope of crRNA design. For our project, a design approach using the AIOD-CRISPR system can be considered in the absence of standard PAM alternatives.
The NUDT_CHINA team proposes: This design also has limitations, requiring that the target sequence is relatively complete, but there may be problems such as fragment breakage in the cfDNA of Echinococcus, and this paper has not attempted to detect multiple sequences in a system. These problems afor us to solve.
In terms of HP, we exchanged the work carried out by both sides in the early stage and developed a science education plan. In the following National Science Popularization Day, CSU_CHINA will be responsible for promotion parts, while NUDT_CHINA responsible for lecturing as well as the experimental parts.
In terms of art, we decided to collaborate on flyer and poster design. As for the video design to exchange ideas and technical details, which could make our work more effective.
In terms of the wiki, the two teams exchanged issues on the construction of the web framework and discussed in detail with the problems arose on both sides, which eventually solved the problem of pipeline failure on NUDT_CHINA's wiki.
3.The specificity test
Initially, we used E gene as a template and different sgRNA to detect amplification products to verify the CRISPR-Cas system.
When we communicated with NUDT_CHINA team, they suggested using different parasite genes (such as Schistosoma japonicum) as templates for amplification to validate our designed sgRNA to verify the specificity of the system.
Indeed, this approach better illustrates the advantages of our system: high specificity and no cross-influence between parasite genes.
Thanks to NUDT_CHINA for their advice, we did the following experiments to optimize our project.
Blind test results of simulated patient samples
Schistosoma japonicum plasmid
The specificity test of the test method indicates that this test method will not detect other parasitic diseases as the positive, nor detect human DNA as the positive, and the primer sequence is specific to the sgRNA sequence. Normal human DNA, Schistosoma DNA and hydatid DNA were blindly tested, and multiple sgRNA-guided cas12a digestion was performed after multiple primer RPA amplification.
4.NUDT_CHINA provides vector expressing CRISPR Cas12a protein
In order to ensure the stability of the experimental results, the CRISPR Cas12a protein purchased from NEB was used in a large range in this project. Although the problems of buffer independent preparation were gradually solved, the protein purchase was still relatively expensive. Although the cost of reagents in the laboratory is more reasonable, our goal is to apply our results to the more remote pastoral areas, and the cost control becomes an urgent problem to be solved.
This year, we focused on exploring and stabilizing the RPA amplification system and the CRISPR Cas12a detection system, without focusing on the plasmid design, so the plan to independently express the CRISPR Cas12a protein was put on hold. Fortunately, the NUDT_CHINA has been focused on building efficient expression systems for many years, and provided us a plasmid sequence expressing CRISPR Cas12a protein efficiently. Obstacles been removed, we could better focus on optimizing our amplification and detection system.
During the subsequent cooperation, the two teams exchanged needs and opinions. The NUDT_CHINA team provided our guidance on designing the highly efficient CRISPR Cas12a protein expression system and designed the expression plasmid to fill in the gaps in our project design.
5.Sep 4th offline meeting up
During the discussion, NUDT_CHINA provided many constructive suggestions for CSU_CHINA, such as presentation and background improvements. NUDT_CHINA mentioned their experience in ethical and legal issues in previous years, which provided great help and reference for CSU_CHINA.
During the communication, we had a very in-depth discussion with NUDT_CHINA about the project and experimental content, such as the source of sgRNA, the noise of cfDNA, and the advantages and disadvantages of RPA. Through in-depth discussion, CSU_CHINA not only found some unnoticed problems, but also gained a lot of novel ideas.
CSU_CHINA introduces the project
The instructor NUDT_CHINA issued suggestions.
As for the NUDT_CHINA project, the two sides also held very in-depth discussions and exchanges. CSU_CHINA has given some meaningful suggestions in the optimization of experimental materials and signal detection. At the same time, with variform HP experience, CSU_CHINA provides many reference examples for NUDT_CHINA members.
6.Sep 25th Science Popularization Day
In the education section, we participated in the National Science Popularization Day together, which aims to popularize the knowledge of synthetic biology to the public.
The content of the activity mainly includes a lecture of synthetic biology and an experiment for primary students. Before the start of the event, we prepared exquisite publicity materials in advance and arranged them in the venue. After the beginning of the activity, we first introduce the basic knowledge of synthetic biology to the masses, and then guide the participants to carry out the experimental operation. Finally, we distributed the brochure of synthetic biology to the people present.
The NUDT_CHINA team is mainly responsible for carrying the materials needed for the experiment, while the CSU_CHINA team is responsible for designing and producing promotional materials. This activity has achieved a very good response, and the people who participated in the event also raised a lot of questions to us, which played a feedback role in the design of our project.
CSU_CHINA laid out promotional props
The members of the two teams work together to guide the students in the experiment.
7.Summary
We have established an in-depth partnership with NUDT_CHINA, whether it is experimentation, modeling, or human practices, and our cooperation in all aspects has enabled both teams to reduce their workload and better complete their projects.
The plasmid map provided by NUDT_CHINA helps us to solve the cost problem, so that we can spend more efforts to improve our amplification-detection system; The offline meeting held with NUDT_CHINA provided an indispensable role for the project of us, which allows us not only to find many problems undiscovered, but also get a different perspective.
We can look at our projects with diversified thinking, which is of great significance for the progress and innovation of our project. From the experience introduced by NUDT_CHINA in previous years, our team has also learned a lot and avoided many problems;The success of the Science Day activities is inseparable from the joint cooperation with NUDT_CHINA. Whether in the planning, preparation or implementation stage of the event, team members on both sides are closely involved and actively follow up.
On the whole, the partnership between the two teams has been very successful this year, and we look forward to further cooperation.
Click here to NUDT_CHINA's Partnership page
