Part:BBa_K817033:Design
PfadBA-RBS-mRFP
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 656
Illegal AgeI site found at 768 - 1000COMPATIBLE WITH RFC[1000]
Design Notes
PfadBA-RFP contains two parts (1)fatty acid-dependent promoter(pfad) and ribosomal binding site(RBS) (2)mRFP, a red fluorescent protein. The promoter pfad acts as a switch of downstream protein expression, and is regulated by the fatty acid-dependent repressor, FadR. mRFP plays a role as a reporter that indicates the expression or efficiency of the promoter.
In the absence of fatty acids, FadR binds at a site downstream of the pfad promoter and represses transcription. When long chain fatty acids become available, they bind to FadR, and elicit a conformational change that releases the protein from DNA, thus removing the repression[1].
In our design, FadR is constitutively expressed and could be repressed by adding oleic acid(a long-chain fatty acid), releasing the FadR and thereby expressing mRFP.
For functional assay result, please refer to our [http://2012.igem.org/Team:NTU-Taida/Result/pFadBA wiki page].
Source
pfad:72bps, from de novo synthesis
RBS:12bps, from BBa_K093005
mRFP:681bps, from BBa_K093005
References
[1] The FadR.DNA complex. Transcriptional control of fatty acid metabolism in Escherichia coli. J Biol Chem. 2001 May 18;276(20):17373-9