摘要
目的 利用脂质体介导法在体外建立转肝细胞生长因子 (HGF)基因的人肝细胞模型。方法 建立HGF真核细胞表达载体 ,利用脂质体介导法在体外将HGF基因转染入人肝细胞 ,利用荧光显微镜观察、免疫组织化学、原位杂交方法检测HGF真核细胞表达载体的转录和表达情况。结果 以阳离子脂质体LipofectAMINE为载体将HGF基因转染人肝细胞后 ,经 40 0mg/L的G41 8筛选后可形成抗性克隆 ;Neo基因原位杂交结果显示转染基因的细胞有阳性表达 ;荧光显微镜下观察到有绿色荧光 ;免疫组织化学证实转染HGF基因的肝细胞有HGF蛋白的表达。结论 HGF基因可被成功转染入人肝细胞并能有效表达 。
Objective To investigate hepatocyte growth factor (HGF) gene transfer and expression in human hepatocyte with LipofectAMINE in vitro.Methods HGF gene was transfected and expressed in human hepatocyte with liposome.The expression of HGF gene was detected by fluorescence microscopy,immunohistochemistry and hybridization in situ.Results After G418 selection,hepatocyte clones were obtained successfully.The positive hepatocytes of in situ hybridization of specific stained Neo probe was observed in hepatocytes transferred eukaryocyte expression vector.Under a fluorescent microscope,the expression of green fluorescent protein (GFP) was seen in hepatocytes transferred vector containing GFP gene.Positive hepatocytes of HGF immunohistochemistry stain was only observed in hepatocytes transferred HGF gene.Conclusion Hepatocyte model transferred HGF gene was successfully constructed.pEGFP HGF expression vector makes it easy to assess the expression of HGF GFP fusion protein and the protein localization on transfected hepatocytes.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2002年第2期152-153,T005,共3页
Chinese Journal of Experimental Surgery
