In Japan, Takashi Ito, professor of genome biology, Cancer Research Institute, Kanazawa University, was reaching the same conclusions. "When we finished the sequencing of the budding yeast, we learned that almost half of the genes within the genome ... hadn't been hit by the genetic approach," he says. That necessitated a new way of doing things: working on the proteins themselves. Not being a protein chemist, Ito says, yeast two-hybrid looked like the way to go.
The premise was simple; the reality was Herculean. This reporter-based assay relies on the dichotomous nature of certain transcriptional activators. One gene of interest can be fused to the DNA binding domain, and another gene believed to interact with the first can be fused to the transcription activation domain. The clones are transformed into haploid yeast, which are then mated. If these proteins interact, the transcription factor becomes functional, switching on expression of the ...
