Team:NEU China B/Collaborations
Collaboration with Fujian normal university
NEU:
Our two teams focused on the problems encountered in the experiment, including some enzymes that we did not use like Gibson enzymes. Our team mainly raised the problem of the actual operation of importing two plasmids into bacteria. In response to this problem, the students of Fujian Normal University gave us a solution.
（photos about our meeting)
We have two teams in the construction of the plasmid also carried out a certain level of cooperation. The students of Fujian Normal University helped us to characterize plasmid p-LsrA-GFP. We helped them to measure the fluorescence value of proU-GFP and construct the plasmid. (http://parts.igem.org/Part:BBa_K2570020)
They tell us that we need to import the first plasmid and then the second plasmid. In addition, they also provide us with some experience about fermentation industry.
Collaboration with SCAU
Our team and the iGEM team of South China Agricultural University conducted an exchange discussion at South China Agricultural University in August 2018. This meeting is mainly to share the two Team's project main content and two teams each encountered in the experiment Problem.
In the meeting, the students of South China Agricultural University show us some their Mathematical models, and we also discuss some problems about our experiments.
Q1: How to match the concentration of AI-2 expressed by the first plasmid pCFDuet-1 with the GFP concentration expressed by pet28a.
A: Our experiment was to release the AI-2 signal molecule after the PCFDUET-1 expression plasmid induction of lactic acid concentration, and AI-2 signal molecule can inhibit the lldr sequence on the second plasmid pet28a.
The LLDR sequence is capable of producing GFP-deterring proteins. In the experiment, we verified that two plasmid copies were more similar, and that the O1o2 sequence could be started normally when the lactic acid concentration was lower (less than 1m mol), and then the GFP gene could be produced fluorescent signal.
Therefore, the expression of two plasmids is more matched, and the experimental results are not affected in this project.
Q2: How do you ensure that your transformant plasmids are not lost?
A: We used the cyanobacterial efficient shuttle plasmid pfq20 given to us by the Wuhan Aquatic Institute of China. We inserted the target gene into the middle of the homologous arm slr0168 at both ends of pfq20. The homology arm can integrate our target fragment into our cyanobacterial genome, allowing our inserted genes to be stably expressed in cyanobacteria, so there is no problem of plasmid loss.
After the meeting, they also give us some help about our experiment. As our project is closely related to lactic acid fermentation, students from South China Agricultural University have provided us with yogurt for the detection of lactic acid content, providing samples for our Experiments.