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Construction of eukaryotic expression vector of humanarresten gene and its secreted expression in hek 293 cells

Frontiers of Medicine 2009年第3期3卷 297-302页

作者： Wei LI Zifang SONG Qichang ZHENG Jun XIONG Dan SHANG Siming GUAN Xiaogang SHU Department of Gerontology Union HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhan 430022China Department of General Surgery Union HospitalTongji Medical CollegeHuazhong University of Science and TechnologyWuhan 430022China

The eukaryotic expression vector of human arresten gene was constructed and its secretive expression in human embryonic kidney(hek 293)cells was *** arresten gene was ampliﬁed from recombinant plasmid pGEM-Arr by polymerase chain reaction(PCR),and then digested with restriction endonucleases BamH I and EcoR *** target fragment was inserted into the BamH I and EcoR I restriction sites of eukaryotic expression vector pSecTag2 to construct ***-tion analysis and DNA sequencing indicated that the arresten gene was successfully inserted into *** recombinant plasmid was subsequently transfected into hek 293 cells with LipofectAMINETM2000 Reagent,and the expression of the target gene was *** transcription PCR(RT-PCR)revealed that the mRNA of the target gene was transcribed in the transfected hek 293 *** blot analysis veriﬁed that the recombinant protein in supernatants was *** supernatants of transfected cells were prepared,and 3-(4,5)-dimethylthiazol(-2-y1)-3,5-di-phenyltetrazoliumbromide(MTT)assay was carried out to assess their effects on the proliferation of human umbilical vein endothelial cells,which showed that the recombinant protein could signiﬁcantly suppress the proliferation of human umbilical vein endothelial cells in *** results provided a solid foundation to explore the usage of arresten in tumor anti-angiogenic gene therapy.

关键词： angiogenesis inhibitor arresten eukaryotic expression hek 293 cells endothelial cells

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