Team:PKU Beijing/Parts Characterization/BBa K228009

Parts > Parts Characterization > BBa_K228009

BBa K228009 - AraC protein(reversed sequence) and Pbad promoter
Designed by Rengcheng Gao Group: iGEM09_PKU_Beijing   (2009-09-18) Input: Arabinose molecules Output: GFP fluorescence

Description
We have constructed the inducement system composed of L-Arabinose inducible Pbad promoter and constitutively expressed AraC regulatory protein. When arabinose is absent, the AraC protein binds to and represses Pbad very efficiently. Thus, the transcription of any downstream coding gene is restricted. On the other hand, the import of arabinose could change the conformation of AraC protein, successfully prevent it repressing Pbad promoter and activate the transcription of downstream coding gene, such as GFP.

This part is constructed for two purposes. First, we coupled two parts, an AraC protein pand a Pbad promoter, in order to characterize the latter one (Part BBa_I13453 ) and obtain more information about this promoter. We place a GFP coding gene(Part BBa_E0840 ) downstream of Pbad promoter to construct a report system. If arabinose is added to the culture, the expression of GFP will be triggered and green fluorescence can be tested. We have tested the function of this promoter according to concentration gradient and time scale, as is shown below. Secondly, we use this part as a sensor in our AND gate system, which works very efficiently.

The direction of AraC coding sequence is opposite compared to the Part BBa_C0080. We have PCRed BBa_C0080 with primers which include standard enzyme-cutting sites. The aim of this reversion is to avoid unexpected expression leaking of downstream sequences.

Safety
Because this part is from commonly used elements in bacteria. No safety problem can be arised by this part.

Compatibility
Chassis: Device has been shown to work in DH5α. Plasmids: Device has been shown to work on pSB1A2 and pSB1A3, pSB4K5 Devices: Device has been shown to work with BBa_K228012, BBa_K228038 - BBa_K228046, BBa_K228110 - BBa_K228118, BBa_K228255 - BBa_K228263, BBa_K228823 - BBa_K228831, BBa_K228870 - BBa_K228878.