Part:BBa_K4239009
toxin/antitoxin gene hok/sok
Description
hok/sok is a toxin/antitoxin system to be used on the same plasmid than an interest gene (the fiatlux operon, in our project). This method of selection ensures that all the bacteria present on the plant have the plasmid, and can hence emit light. Toxins have a longer life span in cells than antitoxins, so if the bacteria lose their plasmid, the toxin is no longer destroyed by the antitoxin. This results in the death of the bacteria, thus assuring the death of all fiatlux non-carriers. As a consequence, by involving a selective pressure, the toxin/antitoxin system increases the stability of the plasmid in the bacteria.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Construction
The gene hok/sok was adapted to fit the biobrick format
Characterization
To check that the synthesized DNA contains the hok/sok toxin/antitoxin system, we carry out a miniprep of the plasmid pSB1C3-hok/sok. This last one is digested by two enzymes, EcoRI and PstI (iGEM restriction sites), and a migration on agarose gel was realized. The expected fragments are :
- Non Digested plasmid : 2664 bp
- Double Digested plasmid : 635 bp + 2030 bp
Column 1: Non digested plasmid. Two strips at 2000 bp and 2600 bp indicates the supercoiled and normal form of the complete non digested plasmid.
Column 2: Double digested plasmid by EcoRI and PstI. The 2 strips at 635 bp and at 2000 bp were expected, and indicate that hok/sok is in the plasmid.
References
Gregor C, Gwosch KC, Sahl SJ, Hell SW. Strongly enhanced bacterial bioluminescence with the ilux operon for single-cell imaging. Proc Natl Acad Sci U S A. 2018 Jan 30;115(5):962-967. doi: 10.1073/pnas.1715946115. Epub 2018 Jan 16. PMID: 29339494; PMCID: PMC5798359.
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