title: Determining the RNA specificity and targets of RNA-binding proteins using a three-hybrid system.
abstract: The three-hybrid system can be used to identify RNA sequences that bind a specific protein by screening a hybrid RNA library with a protein-activation domain fusion as 'bait.' These screens complement biochemical techniques, for example, SELEX, co-immunoprecipitation, and cross-linking experiments (see UV crosslinking of interacting RNA and protein in cultured cells and PAR-CLIP (Photoactivatable Ribonucleoside-Enhanced Crosslinking and Immunoprecipitation): a step-by-step protocol to the transcriptome-wide identification of binding sites of RNA-binding proteins).
authors.affiliation: Microbiology Doctoral Training Program, University of Wisconsin - Madison, Madison, WI, USA.
authors.forename: Yvonne Y
authors.initials: YY
authors.lastname: Koh
authors.affiliation: Department of Biochemistry, University of Wisconsin - Madison, Madison, WI, USA. Electronic address: wickens@biochem.wisc.edu.
authors.forename: Marvin
authors.initials: M
authors.lastname: Wickens
mesh_terms: D015723:Gene Library; D017930:Genes, Reporter; D010957:Plasmids / Q000235:genetics; D011485:Protein Binding; D012313:RNA / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae; D014170:Transformation, Genetic; D020798:Two-Hybrid System Techniques; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
chemical_list: D016601:RNA-Binding Proteins; D012313:RNA; D001616:beta-Galactosidase
keywords: 3-AT concentration; Autoactivation test and sequencing; HIS3 and lacZ activation; Hybrid RNA library; Mutant and control proteins; RNA plasmid; RNA–protein interactions; Three-hybrid system; Transformation efficiency; Z-buffer