Part:BBa_K2631001
BD-ScFv2-pGAL1-eYFP
This construct base on the IGEM parts (BBa_K2247009) contains an anti-AFB1 single chain variable fragments (ScFv2) targeting different affinity sites of AFB1 were fused with DNA binding domain (BD) , and ADH1 terminator (transcription terminator for alcohol dehydrogenase 1). We put another ADH1 terminator, the GAL1 promoter (inducible promoter, regulated by Gal4 in yeast, is used as the promoter in a standard yeast-two-hybrid system) cloned from S. cerevisiae (Johnston M et al, 1984) and an eYFP (enhanced YFP mutated from Aequorea Victoria GFP protein) downstream from the ScFv2 in the BamHI site of the vector (Biteen J S et al, 2008). This part, together with AD-ScFv1 (BBa_K2247008), plays the central role in the AFB1 biosensor system. The presence of AFB1 would draw near the two ScFvs, thus enabling the association of Gal4 AD and BD domains, and activate the EYFP protein, and driving the expression of the reporter gene in the AH109 yeast strain (from Takara). This design is that expression of eYFP under the control of the GAL4 induced promoter it regulates would produce a feedback, making the construct more sensitive to the AFB1 in the environment. eYFP act as a signal of the system are working.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal XhoI site found at 218
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 1457
Illegal AgeI site found at 1539 - 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI site found at 137
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