Team:NCTU Formosa/Basic Part
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Basic Parts
The main goal of our project, E. Phoenix, is to develop a brand new biosystem that can detect mutagens surrounding us. We hope to mutate genes by exposing E. coli to severe surroundings and detect the mutation by toxin gene. This year we decided to design five basic parts. These parts contain sequences of different toxin gene, respectively. They all show the inhibition of bacteria growth. We listed all the basic parts we constructed below.
Toxin Gene
ccdB*(BBa_K3256440)
The original part is from BBa_P1010. The modified amino acid sequence includes N95T. And the protein was truncated at 96th amino acid with a stop codon. After these modifications, the toxin retains the function of suppressing the growth of E. coli, and makes the E. coli more stable from being killed easily.
MazF*(BBa_K3256441)
MazF is toxin protein in MazE-MazF, toxin-antitoxin module. It is function as mRNA endonuclease. The original part is from BBa_K1096002,.The modified amino acid sequence includes P30R, A31R, V32W, V33C. And the protein was truncated at 34th amino acid with a stop codon. After these modifications, the toxin retains the function of suppressing the growth of E. coli, and makes the E. coli more stable from being killed easily.
YafQ*(BBa_K3256442)
YafQ is a toxic gene of the toxin-antitoxin (TA) module in E. coli. YafQ protein will inhibit translation in E. coli. According to researches, gene expression in E. coli decreases 50% after 30 minutes and 70% after 90 minutes since YafQ starts expressing. DinJ is the cognate antitoxin gene of YafQ. DinJ protein will form complex with YafQ to deactivate YafQ. That is, proteins can be synthesized normally once DinJ is expressed. The modified amino acid sequence includes W30C. After these modifications, the toxin retains the function of suppressing the growth of E. coli, and makes the E. coli more stable from being killed easily.
ydfD(BBa_K3256443)
ydfD is one part of the Qin defective prophage in E. coli. According to the research, 99.9% of cell lyses within two hours after ydfD start expressing. dicB, upstream gene of ydfD in E. coli, will prevent bacteria from lysing due to the effect of ydfD. Cell division won’t happen due to inhibit the enzyme that lead to cell division.
ChpBK*(BBa_K3256444)
ChpBK, one of the TA modules in E. coli, is a toxic gene that will suppress cell growth and may cause cell death. Its amino acid sequence holds 35% identity with MazF and 52% similar to it, a famous mRNA interferase in E. coli that can cause cell death. The cognate antitoxin of ChpBK is ChpBI, which can deactivate ChpBK by binding with it. Synthesis of protein can be processed normally once ChpBI is expressed. The modified amino acid sequence includes V83C, R84A. And the protein was truncated at 85th amino acid with a stop codon. After these modifications, the toxin retains the function of suppressing the growth of E. coli, and makes the E. coli more stable from being killed easily.
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