Article

Molecular and Cellular Biochemistry

, Volume 172, Issue 1, pp 67-79

Identification of proteins that interact with a protein of interest: Applications of the yeast two-hybrid system

  • R. Daniel GietzAffiliated withDepartment of Human Genetics, University of Manitoba
  • , Barbara Triggs-RaineAffiliated withDepartment of Biochemistry and Molecular Biology, University of Manitoba
  • , Anne RobbinsAffiliated withDepartment of Human Genetics, University of Manitoba
  • , Kevin C. GrahamAffiliated withDepartment of Human Genetics, University of Manitoba
  • , Robin A. WoodsAffiliated withDepartment of Biology, University of Winnipeg

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Abstract

The yeast two-hybrid system is a molecular genetic test for protein interaction. Here we describe a step by step procedure to screen for proteins that interact with a protein of interest using the two-hybrid system. This process includes, construction and testing of the bait plasmid, screening a plasmid library for interacting fusion proteins, elimination of false positives and deletion analysis of true positives. This procedure is designed to allow investigators to identify proteins and their encoding cDNAs that have a biologically significant interaction with your protein of interest.

protein interaction two-hybrid system yeast screening false positive elimination