【摘要】 目的 构建含HGF基因真核表达载体pEGFP-N1-HGF，观察HGF基因在人脐静脉内皮细胞中表达。为HGF用于血管重建的基因治疗提供实验依据。方法 由重组质粒PBSKSHGF中扩增出HGF基因,将其克隆到含增强型绿色荧光蛋白的真核表达载体EGFP-N1中,应用脂质体将重组质粒EGFP-N1-HGF转染到人脐静脉细胞株ECV304中,G418筛选获得稳定表达克隆,采用荧光显微镜观察、RT-PCR、免疫细胞化学方法鉴定。结果 经G418筛选后可形成抗性细胞克隆；荧光显微镜下观察到有绿色荧光;RT-PCR能扩增出HGFmRNA预期的699bp片段；免疫细胞化学证实转染HGF基因的细胞有HGF蛋白的表达。结论 重组质粒EGFP-N1-HGF能够在细胞株ECV304转录、表达。

    【关键词】 HGF基因;内皮细胞；EGFP;

    Construction of EGFP labeled recombinant vector containing HGF and its expression in vein endothelial cells

    CHANG Bing-mei,LI Mei-ning,XIE Jun,YANG Tao,TIAN Xin-rui,CHENG Niu-liang,NIU Bo.

    Biochemistry & Molecular Biology Department of Shanxi Medical University,Taiyuan 030001,China

    【Abstract】 Objective To investigate hepatocyte growth factor (HGF) gene transfection and expression in human umbilical vein endothelial cells( ECV304) with liposome in vitro.Methods The recombinant plasmid was constructed and transfected into human umbilical vein endothelial cells with liposome：GeneSHUTTLE-20, and positive clones were selected by G418. The transfection and the expression of HGF in ECV304 were tested by RT-PCR and immunohistochemistry.Results After G418 selection, the cell clones were obtained successfully. Under a fluorescent microscope, the expression of enhanced green fluorescent protein(EGFP) was seen in cells transfected with vector contain EGFP gene. RT-PCR and immunohistochemistry proved that there was transcription of HGF gene in transfection cells and there was expression of HGF in these cells.Conclusion The recombinant of EGFP-N1-HGF is an effective expression vector. The recombinant of EGFP- HGF transcript and express in the transfected cells. ......