Part:BBa_K729008

pCstA+RBS+T7RNAP+pT7+RBS+GFP+TT

This starvation sensitive device was used by the [http://2012.igem.org/Team:University_College_London UCL’s 2012 iGem team] to increase the expression of a reporter gene (GFP) under low glucose concentrations. During carbon starvation cAMP binds the cAMP receptor protein, the complex formed activates the expression of the downstream gene (T7 RNA polymerase), which promotes the expression of GFP under the control of the T7 promoter.

This device comprises the ligation of the parts (BBa_K729007) and (BBa_I746909).

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 2793
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 2887
  

Characterisation

Normalized expression levels were plotted as relative fluorescent units per optical density at 600 nm. Transformed E. coli was grown overnight in LB media and then transferred into minimal M9 media supplemented with different glucose concentrations (0%, 0.2%, 0.5%, 1.0%, 3%, 5%) to induce the activation of the cstA promoter. BBa_K729008 integrates the T7 RNA polymerase and T7 promoter in order to increase the expression of the reporter gene. Once the bacteria is exposed to carbon starvation stress, cstA promoter is activated, as a results it can be seen from the chart that GFP was expressed reaching higher accumulation in cultures with M9 media supplemented with less than 0.5% of glucose.