From 2014.igem.org

Biology - Classic & RNAseq-based sensors

Phenol biosensor :

PCBs biosensor :

• Biosensor parts
  
  Pseudomonas pseudoalcaligenes KF707 is one of the strain which are able to degrade polychlorinated biphenyls (PCBs). The bacteria contains a biphenyl-catabolic (bph) gene cluster (bphR1A1A2-(orf3)-bphA3A4BCX0X1X2X3D). bphR1 is a positive regulator for itself and bphX0X1X2X3D and is responsible of the degradation of PCBs. Watanabe et al showed that an other regulatory protein, bphr2, is involved in the regulation of theses genes. bphR2 can detect PCBs when it diffuses into the cell and activate degradation genes by activating the transcription of bphR1.
  .
  
  In 2013, Saclays' team wanted to construct a biosensor for PCBs, a project that failed. So our first aim was to finish their construct and to optimize it. The construction is composed by a constitutive promoter (BBa_J23114), a RBS (BBa_B0034), bphR2 gene (BBa_K1413021), which has been mutated because of a pstI site in its sequence, bphR1 promoter region (BBa_K1155001), received from Saclay’s team, RFP (BBa_E1010) and a terminator (BBa_B0015)
  
  
  
  
  
  
• How function our biosensor ?
  
  In absence of PCBs :
  bphR2 (BBa_K1413021) is bound to bphR1 promoter (BBa_K1155001). Transcription of RFP isn't possible.
  
  
  
  
  In presence of PCBs :
  When compound diffuses into the cell, it binds to bphr2 protein. This protein undergoes a conformational change and releases from the promoter that its allows the transcription of RFP.
  

• Improvement
  
  Their first problem was a pstI site in their bphR2 sequence (BBa_K1155009) so this gene has been mutated by keeping the same codon bla bla bla