| The two-hybrid system is a molecular genetic test in Saccharomyces cerevisiae that can identify and study protein:protein interactions in vivo. This system can be used to identify the motifs that are involved in the interaction of two proteins. Alternatively, the system can be used to identify novel binding partners of a protein of interest. Currently, all yeast strains used for the two-hybrid system have numerous features which prevent full utilization of the system. This thesis describes the construction of two new and improved yeast strains for use in the two-hybrid system. The yeast genes URA3, LYS2 and LEU2 were manipulated to form three disruption cassettes. These cassettes were used to disrupt the URA3 and LYS2 genes and stably integrate GAL1-lacZ and GAL1-HIS3 reporter genes. Both reporter gene constructs have low levels of basal activity allowing for the detection of weak protein:protein interactions. The strain KGY37 can be used with Gal4... |
