Part:BBa_K200018

pCstA+RBS+GFP+TT

This BioBrick comprises a ligation of the registry parts for the promoter PcstA (BBa_K118011) and the RBS (BBa_B0034) to GFP (BBa_E0040) and a double terminator (BBa_B0015).

The PcstA promoter is cAMP activated. Under low glucose concentrations, there is increased activity by adenylate cyclase. This results in cAMP binding to the cAMP receptor protein, and activating the promoter for downstream expression (more information on this part can be found on its registry page here).

This part contains a GFP (green fluorescent protein) reporter in the functional transcriptional unit. As such, activity of the pCstA promoter can be assessed by the expression of GFP.

Usage and Biology

This construct was used by Imperial College's 2009 iGEM team for [http://2009.igem.org/Team:Imperial_College_London The E.ncapsulator] project using the following subparts: BBa_K118011, BBa_B0034, BBa_E0040 and BBa_B0015. It was ligated to BBa_J5526 to produce the final construct BBa_K200019.

Characterisation

Cells with BBa_K200018 were grown overnight in various different media, and the GFP fluorescence was measured.

After overnight culture, the corrected fluorescence of glucose is almost negligible, showing that glucose represses the PcstA promoter strongly.

For all the other secondary carbon sources, 10% Casamino Acids in M9 shows the highest corrected fluorescence at 22000.

For information, go to the Imperial iGEM 2009 E.ncapsulator project page on [http://2009.igem.org/Team:Imperial_College_London/Wetlab/Results/CRP_and_Media|BBa_K200018 testing]





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
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 801