title: Detecting protein-DNA interactions using a modified yeast one-hybrid system.
abstract: The yeast one-hybrid (Y1H) system has been among the methods of choice to detect protein-DNA interactions. However, conventional Y1H systems with a single auxotrophic reporter gene often suffer from high incidence of false positives to demonstrate a limited power in large-scale screenings. Here we describe a refined Y1H system that uses two independent bait sequences, each controlling a distinct reporter gene integrated in the host genome. With these modifications and a method of targeted DNA methylation, we succeeded in efficient isolation of clones for methylated DNA-binding proteins from mammalian cDNA libraries.
authors.affiliation: Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo, 113-0032, Japan, oota@bi.s.u-tokyo.ac.jp.
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Ota
authors.forename: Shu-Ying
authors.initials: SY
authors.lastname: Feng
authors.forename: Takashi
authors.initials: T
authors.lastname: Ito
mesh_terms: D000818:Animals; D001483:Base Sequence; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D019175:DNA Methylation; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015723:Gene Library; D017930:Genes, Reporter; D006801:Humans; D010957:Plasmids / Q000235:genetics; D014170:Transformation, Genetic; D020798:Two-Hybrid System Techniques; D015003:Yeasts / Q000235:genetics / Q000378:metabolism
chemical_list: D004268:DNA-Binding Proteins; D004247:DNA