Construction and identification of recombinant adenovirus vector containing human ?-catenin / 重庆医科大学学报

ABSTRACT

Objective:To construct the recombinant adenovirus vector containing human?-catenin gene by homogenous recombination in E.coli for later research of the mechanism of Wnt/Wingless signal pathway in tumorigenesis.Methods:The target gene was amplified from pcmv-?NS-FC plasmid by PCR,then cutted by two endonucleases and directional cloned to the pAd track-cmv vector.Afterward,the correct recombinant was linearized by PmeI,following co-transformation with the backbone vector pAd Easy-1 in E.coli BJ5183.The homologous recombinant was linearized by PacI,and then transfected into HEK293 cell line to pack the adenovirus.Results:Through PCR,endonuclease cutting and gene sequencing,the target gene was verified to be correctly cloned in adenovirus vector.The high expression of green fluorescence protein in 293 cell line was found under fluorescent microscope.Conclusion:The recombinant adenovirus vector containing human ?-catenin gene was successfully constructed and the adenovirus was packaged in HEK293 cell line.