Part:BBa_K137117

AHL induced expression of rcsA with constitutive cI

The strong AHL inducible promoter upstream of rcsA as the inducible switch for activating the lytic cycle. The constitutively active cI is for reducing activation due to leaky rcsA expression.

Usage and Biology

The phage λcam was used to establish lysogens of E. coli strain DH10B carrying rcsA induction devices in pSB1A2. Using the Caltech iGEM [http://2008.igem.org/Team:Caltech/Protocols/rcsA_Lysogen_Induction rcsA lysogen induction protocol] and then [http://2008.igem.org/Team:Caltech/Protocols/Titering titering] the resulting phage on wild-type E. coli, the series of devices S04080 (no promoter), S04087 (weak RBS B0033), K137118 (medium RBS B0032), K137114 (strong RBS B0034), and K137117 (strong RBS B0034 plus constitutive cI expression) produced the following concentrations of plaque forming units (pfu/mL).

Increasing the RBS strength increased the background lysogen induction without changing the dynamic range (ratio of pfu/mL induced vs. uninduced). Adding constitutive cI expression increased the dynamic range roughly 3-fold, lowering background leakiness without significantly lowering the induced expression.

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 1864
  Illegal NheI site found at 1887
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1005