Part:BBa_K4194012

p(TetR)-RBS-EGFP-LVA-RBS-δ activator-LVA-terminator-pF-RBS-mRFP1-LVA-RBS-TetR-LVA-terminator

This is a gene oscillator. The feedback loop of this gene oscillator consists of pF promoter (activated by δ activator) and Tet gene expression regulation system. δ activator is an activator from phiR73 phage, which can act on a class of inducible promoters including pF promoter. The pF promoter in this system comes from the P2 phage genome. Without activation, the pF promoter can hardly start the expression of downstream genes. When δ activator exists, pF is activated, and the downstream genes are expressed. Julien et al. fused each of P2 and P4 late promoters to lacZ, and analyzed the fusion of each promoter with each activator. By detecting the activity of β-galactosidase, the range of dynamic regulation was determined. Without activator, these promoters have little activity. In the presence of any activator, transcription is induced. The δ activator has a wide range of dynamic regulation of pF promoter, so it is suitable to be a part of the feedback loop. In addition, the promoter of δ activator to pF comes from phage, and it is not easily interfered by other metabolic activities of E.coli, which can ensure the good orthogonality of gene oscillator system. The expression of δ activator is regulated by p(tetR) promoter, which can be repressed by TetR protein. In addition, in order to make the protein degrade in time and make the feedback response more sensitive, ssrA-LVA tag is added at the end of GFP, δ activator, RFP and tetR.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal XbaI site found at 1387
  Illegal PstI site found at 961
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 961
  Illegal NotI site found at 2105
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal XbaI site found at 1387
  Illegal PstI site found at 961
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal XbaI site found at 1387
  Illegal PstI site found at 961
  Illegal NgoMIV site found at 1056
  Illegal NgoMIV site found at 1071
  Illegal AgeI site found at 1314
  Illegal AgeI site found at 1987
  
• 1000
  COMPATIBLE WITH RFC[1000]