Construction of Eukaryotic Expression Vector for Glypican-3 and Its Expression in Cell

doi:10.13560/j.cnki.biotech.bull.1985.2016.05.019

Abstract

Abstract: This work aims to construct the eukaryotic expression vector of glypican-3（GPC3）for acquiring anti-GPC3 monoclonal antibody. The GPC3 gene was amplified by PCR and was cloned into an expression vector p3XFLAG-CMV-14，and the expression vector pCMV-gpc3 was constructed. HEK293 cells were transfected with the vector by liposome，and the final results were detected by Western blot. The stably-expressed cells were collected and ground，and the high-purity GPC3 protein was obtained by affinity column. The eukaryotic expression vector of pCMV-gpc3 was constructed successfully. After the transfection to HEK293 cells，the monoclonal cell line of stably-expressed was gained by G418 screening. Western blot analysis revealed that the target protein expressed markedly. A large number of GPC3 proteins were obtained via genetic engineering and purifying technology，which laid a foundation for the biological function and application of the protein as well as the preparation of monoclonal antibody.

Key words: glypican-3, eukaryotic expression, liposome transfection, affinity purification

CHENG Hua,FANG Xiang-dong,CUI Shuo,WU Meng,ZHANG Zhao-jun,LI Ze-xia. Construction of Eukaryotic Expression Vector for Glypican-3 and Its Expression in Cell[J]. Biotechnology Bulletin, 2016, 32(5): 146-150.

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