Difference between revisions of "Part:BBa K3482004"
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E.coli Nissle 1917 were transformed with a plasmid containing the part BBa K3482004 and the part <html><a style="padding: 0px; margin: 0px;" href="https://parts.igem.org/Part:BBa_K3482014"> BBa_K3482014 </a></html> (IM2 antitoxin part) and plated with a gradient of aTc and IPTG on agar plate. | E.coli Nissle 1917 were transformed with a plasmid containing the part BBa K3482004 and the part <html><a style="padding: 0px; margin: 0px;" href="https://parts.igem.org/Part:BBa_K3482014"> BBa_K3482014 </a></html> (IM2 antitoxin part) and plated with a gradient of aTc and IPTG on agar plate. | ||
The plate shows strong activity of the IM2 antitoxin with aTc induction, whereas IPTG induction promotes production of the MiniColicin E2 toxin, resulting in a number of surviving cells (probable mutants) proportional to the dilution of the plated culture. | The plate shows strong activity of the IM2 antitoxin with aTc induction, whereas IPTG induction promotes production of the MiniColicin E2 toxin, resulting in a number of surviving cells (probable mutants) proportional to the dilution of the plated culture. | ||
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Revision as of 19:57, 27 October 2020
IPTG-inducible miniColicin toxin
This part allows for IPTG inducible expression of the DNAase miniColicin E2 in E. coli. The degradation of DNA , which leads to cell death, can therefore be induced by IPTG.
Biology
Colicin E2 is an endonuclease that cuts on both single and double-stranded DNA. It has no specific cutting site. Colicins are toxins that can be produced by E.coli and closely related bacteria.
Usage
We used that part to test if the expression of the toxin work in our E.coli Nissle 1917 strain and if it was sufficient do provoc cell death
Characterization
Functional kill switch assay with IPTG and aTc gradients on agar plate
E.coli Nissle 1917 were transformed with a plasmid containing the part BBa K3482004 and the part BBa_K3482014 (IM2 antitoxin part) and plated with a gradient of aTc and IPTG on agar plate. The plate shows strong activity of the IM2 antitoxin with aTc induction, whereas IPTG induction promotes production of the MiniColicin E2 toxin, resulting in a number of surviving cells (probable mutants) proportional to the dilution of the plated culture.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 124
- 1000COMPATIBLE WITH RFC[1000]