加强型黄色荧光蛋白和VEGF双基因载体在肝细胞共转染表达

Co-transfection and Expression of Enhanced Yellow Fluorescent Protein and Vascular Endothelial Growth Factor Double Gene Vector on Hepatocyte

【目的】构建携带加强型黄色荧光素蛋白 (EYFP)和人类血管内皮生长因子 (VEGF)的双基因真核表达载体pIRES EYFP/VEGF12 1,用EYFP作标记基因 ,研究外源性VEGF12 1基因在大鼠原代培养肝细胞转染表达 ,为实时监测VEGF12 1基因修饰肝细胞移植后状态提供基础。【方法】将 pcDNA3VEGF12 1中VEGF12 1定向克隆到 pIRES EYFP ,构建重组质粒 pIRES EYFP/VEGF12 1,经酶切、PCR扩增和部分DNA序列分析证实后 ,转染大鼠原代培养肝细胞 ,用荧光显微镜等观察转染表达。【结果】酶切、PCR扩增和部分DNA序列分析等证明pIRES EYFP/VEGF12 1质粒成功构建 ,转染大鼠原代培养肝细胞后 ,可以通过荧光显微镜观察到黄绿色荧光。【结论】构建了 pIRES EYFP/VEGF12 1质粒 ,为实时监测外源性VEGF12 1基因转染表达和VEGF基因修饰肝细胞的基因治疗奠定基础。

To construct double gene eukaryotic expression vector pIRES EYFP/VEGF 121 . By using enhanced yellow fluorescent protein (EYFP) as a marker gene to study the transfection and expression of VEGF 121 gene. VEGF 121 gene was cloned into plasmid pIRES EYFP pIRES EYFP/VEGF 121 was constructed and identified by restriction endonuclease enzyme analysis, PCR amplification and partial DNA sequence analysis Then the plasmid was transfected into primary cultured hepatocyte Expression of exogenous genes were observed through fluorescence microscope pIRES EYFP/VEGF 121 plasmid was identified though restriction endonuclease enzyme analysis,PCR amplification and partial DNA sequence analysis After transfected into rat primary cultured hepatocyte, yellow green fluorescence was observed under fluorescence microscope [Conclusion] The eukaryotic expression plasmid pIRES EYFP/VEGF 121 has been constructed successfully