Part:BBa_K1767003

P(Lac)IQ RBS Aiia RBS LuxR ter ter luxpR RBS tetR ter ter P(tetR) RBS mRFP1 ter ter

We designed and built a kill switch mechanism that based on the activation of a lethal gene downstream of an inducible promoter that is activated over time.

This is the full construct of our genetic circuit that consists of a constitutive promoter (I14032) driving expression of Aiia and LuxR followed by a Lux promoter(R0062). This promoter is activated by the LuxR-AHL complex , driving expression of TetR , followed by a Tet promoter (R0040) . This promoter is repressed by the TetR protein which in absence of TetR, drives expression of RFP.

You can use this BioBreak in several ways: 1.To retrieve information about AHL: in absence of AHL , RFP is expressed and no RFP is expressed in presence of AHL. 2.You can use it as a kill switch, but in order to do that RFP needs to be exchanged with a toxic protein of your choice. If there is AHL added into the medium containing your engineered bacteria, they will stay alive. Over time, AHL is degraded by AiiA (whose activity can be controlled by changing various factors such as the RBS strength and half-life), and the bacteria will die. The scheme below explains how this BioBrick works:

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
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 157
  Illegal AgeI site found at 3309
  Illegal AgeI site found at 3421
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1761