Team
Safety
Awards
Parts
DAY1/2023.7.31
We
shared and discussed the content of the iGEM program and received
training on laboratory safety. Then, we prepared LB liquid medium and
LB solid medium.
DAY2/2023.8.1
We learned the basic principles and procedures of molecular cloning, as well as the use of Snapgene software.
DAY3/2023.8.2
We learned the principles and operations of plasmid extraction and extracted the pET28a-mCherry and pETM6-pnar plasmids.
DAY4/2023.8.3
We double-digested pET28a-mCherry and pETM6-pnar with XhoI and AvrII restriction enzymes, respectively. Then, we performed agarose gel electrophoresis to verify the fragments.
DAY5/2023.8.4
We learned the principles and operations of gel extraction and fragment recovery of the cut strips.
DAY6/2023.8.5
After obtaining the target fragment, we ligated the two fragments using T4 ligase, an enzyme ligation reaction that needs to be performed overnight.
DAY7/2023.8.6
We transformed the recombination product into the E. coli DH5α competent cells, performed a plate-coating operation, and incubated the plates overnight.
DAY8/2023.8.7
Since yesterday's experiment failed, the steps of yesterday's experiment were repeated today.
DAY9/2023.8.8
We identified the transformants by colony PCR and inoculated the transformants with the correct bands.
DAY10/2023.8.9
We used the kit to extract plasmids from overnight cultures for later construction of the RBS mutant library.
DAY11/2023.8.10
We learned how to design mutants for RBS with the RBS calculator (https://salislab.net/software/). Primers with mutant sites were designed for subsequent whole plasmid PCR.
DAY12/2023.8.11
Today, we obtained the RBS1-RBS8 fragments by whole plasmid PCR, blunted the ends of the products, and ligated them overnight with T4 enzyme, respectively.
DAY13/2023.8.12
We transformed the recombination product into E. coli BL21(DE3) competent cells, spread the plates, and incubated the plates overnight.
DAY14/2023.8.13
We identified the transformants by colony PCR and inoculated the transformants with the correct bands.
DAY15/2023.8.14
We inoculated the transformants verified by colony PCR for tomorrow's fluorescence intensity test.
DAY16/2023.8.15
We performed fluorescence intensity tests on the obtained RBS mutant library to determine which RBS sequences have higher translation efficiency.
Organize experimental results and write wiki materials.
