View source for Team:Pitt/3-hybrid/Project

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<h1>VEGF-A Sensor</h1>
<p>This project uses a 3-hybrid approach to activate transcription of a reporter gene based on the presence of VEGF-A. The &alpha;-subunit of E. coli RNA polymerase often recruits the polymerase to the appropriate DNA through its C-terminal domain. By replacing the C-terminal domain with an antibody fragment - known as single chain variable fragment (scFv) - specific to VEGF-A, the &alpha;-subunit will recruit the polymerase to a molecule of VEGF-A. By creating another fusion protein that consists of a DNA-binding protein and another scFv, the same VEGF-A molecule can be recruited to the DNA, thus completing the 3-hybrid system, and allowing the polymerase to transcribe the reporter. While this project is specific to VEGF-A, the system is incredibly easy to modify to work for other similarly-sized biological molecules. By choosing the scFv's used, and the spacing between the upstream promoter element and the transcriptional start site, the specificity of the system is easily customized. This method can potentially replace ELISA in some instances as a faster, cheaper, and more transportable alternative. </p>
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