Modified Yeast-Two-Hybrid System to Identify Proteins Interacting with the Growth Factor Progranulin

Qing-Yun Tian; Yun-Peng Zhao; Chuan-ju Liu

doi:10.3791/3562

Modified Yeast-Two-Hybrid System to Identify Proteins Interacting with the Growth Factor Progranulin

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Biology

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JoVE Journal Biology

Modified Yeast-Two-Hybrid System to Identify Proteins Interacting with the Growth Factor Progranulin

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07:56 min

January 17, 2012

DOI: 10.3791/3562-v

Qing-Yun Tian¹, Yun-Peng Zhao¹, Chuan-ju Liu^1,2

¹Department of Orthopaedic Surgery,NYU Hospital for Joint Diseases, ²Department of Cell Biology,New York University School of Medicine

We have modified the conventional yeast two-hybrid screening, an effective genetic tool in identifying protein interaction. This modification markedly shortens the process, reduces the workload, and most importantly, reduces the number of false positives. In addition, this approach is reproducible and reliable.

This MY two H system improves on the conventional yeast two hybrid protocol to markedly reduce the number of false positives. First, prepare a small scale transformation of the bait plasmid into yeast. After validating the bait construct using a competitive inhibitor screen the cDNA library on minus four plates.

Proceed to verify positive clones with XGL and reation assays. Then identify the potential interacting proteins by DNA sequencing and bioinformatics analysis. Ultimately, this modified yeast two hybrid system significantly increases efficacy and success of the screen process.

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