Interaction mating methods in two-hybrid systems.

Introduction The yeast two-hybrid system is a powerful assay for protein-protein interactions 1. As described elsewhere in this volume, several versions of the twohybrid system have been developed. Most versions have the following features. The two proteins to be tested for interaction are expressed as hybrids in the nucleus of a yeast cell. One of the proteins is fused to the DNA binding domain (DBD) of a transcription factor and the other is fused to a transcription activation domain (AD). If the two hybrid proteins interact, they reconstitute a functional transcription factor which activates one or more reporter genes that contain binding sites for the DBD. This simple assay has been widely used to identify new interacting proteins from libraries, to test interactions between small and large sets of proteins, to map protein networks, and to address the functions of individual proteins and protein interactions 2-5. Here we describe interaction mating, a two-hybrid variation that can be adapted to most versions of the system and which can simplify and facilitate most two-hybrid experiments 6,7.