Part:BBa_K4307005

OR2-OR3-cI + pBAD-cro

cI/Cro is a natural feedback system in bacteriophages, which acts as a bidirectional switch to determine the fate of bacteriophages and the infected bacteria. Phage genome has two operators, called OR (the right operator) and OL (the left operator). OR plays an important role in the regulation of gene expression of phage. The cI protein is a phage-encoded repressor that binds to the OR. After binding to OR, cI protein will exclude RNA polymerase from binding to PR (located within OR) and inhibit the expression of the downstream gene. Cro protein is another phage-expressed protein, which can relieve the inhibition by preventing the synthesis of the repressor.

Considering that the phage genome can be integrated into the bacterial genome in lysogenic bacteria, we isolated the the part related to the expression of cI (OR2-OR3) in OR and integrated it into the bacterial genome together with the downstream cI gene. The Cro gene in phage was isolated and integrated with pBAD promoter. Thus, we successfully reproduced the bidirectional switch in bacteriophage using OR2-OR3-CI and PBAD-CRO in Escherichia coli.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal EcoRI site found at 366
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal EcoRI site found at 366
  Illegal NheI site found at 355
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal EcoRI site found at 366
  Illegal BglII site found at 437
  Illegal BamHI site found at 294
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal EcoRI site found at 366
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal EcoRI site found at 366
  Illegal AgeI site found at 129
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI site found at 111