An efficient method for the isolation of interaction-null/impaired mutants using the yeast two-hybrid technique

Protein-protein interactions are one of the most basic and critical processes underlying biological functions. Thus, identification of the interacting proteins of a protein of interest and further elucidation of the roles of the interactions is critical for understanding the related biological processes. The yeast two-hybrid (Y2H) method is a popular approach for identifying protein-protein interactions. Once interacting proteins are identified, a comparison of the phenotypes of mutants lacking the specific protein-protein interaction with those of the wild-type strain is a powerful tool for uncovering the former interactions' biological significance. However, isolation of such interaction-defective mutants is often laborious. Here, I describe a novel and efficient approach for isolating such mutants that uses the Y2H technique with a modified Y2H vector, and provide an example of how this approach can be used to screen interaction-null/impaired mutants. Because the strategy is simple and the modification of a pre-existing Y2H vector is sufficient for the screening purpose, the same strategy can be applied to any existing two-hybrid system.