Part:BBa_K1075031

AraC-pBAD-RBS32-ccdB-TT

All ccdb parts have been withdrawn from the Registry. Samples of parts containing the ccdB gene cannot be requested. - iGEM HQ

The part contains a chemically induced kill-switch. When Arabinose is added to the bacteria, the toxin ccdB is expressed and cell death is initiated.

Usage and Biology

The ccd module is a toxin-antitoxin (TA) system. The module naturally occurs on the F Plasmid in Escherichia coli bacteria and is essential for their survival. Normally the toxin ccdB is inactivated by the presence of the antitoxin ccdA in the form of a ccdAB complex. If ccdA is no longer available, ccdB inhibits DNA gyrase which leads to cell death. Gyrase is a type IIA topoisomerase specific to E. coli and is able to produce negative DNA supercoiling by making a double-strand break in the DNA and religating it. CcdB stabilizes the gyrase cleavage complex and thus blocks the catalytic function of the gyrase. That means that the gyrase remains bound to the DNA and the cleaved DNA is not religated. DNA- and RNA polymerases can’t copy the DNA anymore and cell proliferation as well as protein biosynthesis is stopped. The double-stranded breaks in the DNA initiate cell death. [http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1635281/][http://www.ncbi.nlm.nih.gov/pubmed/?term=A+Common+Origin+for+the+Bacterial+Toxin-Antitoxin+Systems+parD+and+ccd]

The part was designed for proof of principle of the toxic effect of the toxin ccdB.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 1144
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 979
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1452
  Illegal SapI site found at 961