VEGF-C真核表达载体的构建及蛋白表达

目的构建人血管内皮生长因子(VEGF-C)编码序列基因真核表达载体,为探讨VEGF-C的生物学功能奠定基础.方法根据已公布的序列设计引物,用PCR方法从肿瘤细胞株cDNA中扩增出人VEGF-C编码序列基因片段,测序正确后用限制性内切酶将目的片段插入PcDNA3.1/V5-His-TOPO载体中.采用酶切和PCR鉴定后通过脂质体介导转染至CHO细胞中进行瞬时表达.结论经RT-PCR扩增转染细胞cDNA和Western blotting检测证实重组质粒pcDNA3.1-VEGF-C能在宿主细胞中高效表达.

The Construction and Expression of Eukaryotic Expression Vector Harboring Homo Sapiens Vascular Endothelial Growth Factor-C Gene

Objective To study the biological activity of VEGF-C,and to consbuct eukaryotic expression vectors harboring homo sapiens VEGF-C gene was constructed.Methods The human VEGF-C cDNA was amplied from the pancreatic cancer cell line A549 by PCR and the PCR products which was in line with the reported human VEGF-C gene were inserted into eukaryotic expression vectors PcDNA3.1/V5-His-TOPO by enzyme restriction and ligation. Methods The recombinant expression plasmid was transfected into CHO cells and the transient expression product was analyzed by Western blotting. Conclusion The results demonstrated that the eukaryotic expression vector PcDNA3.1/V5-His-TOPO-VEGF-C was successfully constructed and can express its corresponding protein efficiently, which provides a basis for the further study of biological function of human VEGF-C.