人血管内皮生长因子基因的克隆及真核表达质粒的构建

目的：克隆人血管内皮生长因子基因VEGF165片段。构建pcDNA3／VEGF165真核表达质粒。方法：采用逆转录聚合酶链式反应(RT-PCR)技术，从人肝癌细胞株SMMC-7721中扩增出血管内皮生长因子VEGF165基因片段，通过DNA重组技术将该基因片段重组于pcDNA3真核表达载体上，构建成pcDNA3／VEGF165重组质粒，分别用PCR扩增、酶切电泳分析及DNA测序的方法对重组DNA进行鉴定。结果：经PCR扩增、酶切电泳分析和DNA测序证实，本实验构建的重组质粒目的基因片段为人VEGF165cDNA。结论：本实验成功地克隆了VEGF165基因并构建了其真核表达质粒。为进一步研究用VEGF基因转染的方法来恢复或增强放疗后组织的血管生成能力奠定了基础。

CLONING OF HUMAN VASCULAR ENDOTHELIAL GROWTH FACTOR GENE AND THE CONSTRUCTION OF ITS EXPRESSIVE PLASIMAD

Objective:The aim of this study is to clone VEGF gene and to construct the expressive plamsid pcDNA 3/VEGF 165Methods:Human vascular endothelial growth factor gene was amplified by RT-PCR method from human cells SMMC-7721 strains and constructed into the expressive plamsid pcDNA 3/VEGF 165. The gene in the expressive plamsid pcDNA 3/VEGF 165 is identified by PCR amplifcation, enzyme digestion and DNA sequencing.Result:The cloned DNA was confirmed to be VEGF 165 gene.Conclusion:In this study we successfully cloned VEGF 165 gene and constructed its expressive plamsid pcDNA 3/VEGF 165, which provided the foundation of using VEGF gene therapy to resume or build up the angiogenesis of ischemia tissues in vivo.