Part:BBa_K3482016

IPTG-inducible ccdb toxin

This part allows for IPTG inducible expression of the CcdB toxin in E. coli which leads to cell death by acting on the DNA gyrase. The action of CcdB leads to the degradation of DNA , which results in cell death. The CcdB toxin tagets the GyrA subunit of the DNA gyrase. The CcdB toxin blocks the GyrA subunit and leads to a break into the double stranded DNA.

This part will be used in a kill-switch system with anohter part containing the antitoxin BBa_K3482020 .

Characterization

Functional killswitch assay with IPTG and aTc gradients on agar plate

E.coli Nissle 1917 were transformed with a plasmid containing the part BBa K3482016 and the part BBa_K3482020 (CcdA antitoxin part) and plated with a gradient of aTc and IPTG on agar plate. The plate shows strong activity of the CcdA antitoxin with aTc induction, whereas IPTG induction promotes production of the CcdB toxin, resulting in a number of surviving cells (probable mutants) proportional to the dilution of the plated culture.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 307