Screening of interferonαbinding proteins by yeast-two hybrid technique

Objective To screen proteins binding with interferonα(IFNα)from human hepatic cDNA libraty by yeast-two hybrid technique.Methods The IFNαgene was amplified by polymerase chain reaction(PCR)and constructed into pGBKT7 vector as the bait plasmid in yeast-two hybrid system3,pGBKT7-IFNαwas then transfected into yeast AH109.The transfected yeast were mated with yeast Y187 containing liver cDNA library plasmid in 2×YPDA medium.Diploid yeast was plated on synthetic dropout nutrient medium(SD/-Trp Leu-His-Ade)and synthetic dropout nutrient medium(SD/-Trp-Leu-His-Ade)containing X-α-gal for selecting.After plasmid extracting and en- zyme cutting analysis,the blue colonies were performed sequence analysis,the results were analyzed by bioinformatics.Results IFNαgene was successfully cloned and expressed in yeast cells.Thirty- four positive colonies were obtained using yeast-two hybrid technique.After sequence analysis,eight clones were found may have a binding effect with IFN protein.Conclusions IFNαgenes was success- ful cloned and eight proteins that could bind with IFNαprotein were also screened.