摘要
目的:研究原代培养大鼠肝细胞的基因转染的方法方法:采用胶原酶灌注法获取原代培养大鼠肝细胞.利用脂质体转染法将含有绿色荧光蛋白(GFP)和Neo基因的真核细胞表达载体(pEGFP-N3)转染原代培养大鼠肝细胞.用荧光显微镜观察和Neo基因原位杂交染色方法检测肝细胞内基因表达情况. 结果:获取的原代大鼠肝细胞活细胞率达95%.荧光显微镜下观察可见转染基因的细胞可发出绿色荧光,原位杂交显示有Neo基因的表达. 结论:pEGFP-N3基因可转入大鼠肝细胞并获得表达,可用于标记原代培养的大鼠肝细胞,有利于研究肝细胞移植后移植的肝细胞在体内的分布及功能.
AIM: To study the efficient and stable gene transfection method of rat primary cultured hepatocytes by liposome. METHODS: Rat hepatocytes were isolated by collegenase perfusion, and the pEGFP-N3 plasmid containing GFP and Neo gene was transfected into rat primary hepatocyte with liposome. The expression of GFP and Neo was observed by fluorescence microscopy and in situ hybridization. RESULTS: The rate of live hepatocytes obtained was 95%. Under fluorescence microscope, hepatocyte transfected with pEGFP-N3 plasmid showed green fluorescence. By using in situ hybridization method, expression of Neo gene was observed in hetpatocyte transfected with pEGFP-N3. CONCLUSION: Rat hepatocyte can be transfected by pEGFP-N3. pEGFP-N3 expression vector and makes it easy to assess the expression of target gene in transfected hepatocyte, and it can be used for localization of transplantated hepatocyte.
出处
《世界华人消化杂志》
CAS
2004年第3期642-645,共4页
World Chinese Journal of Digestology
基金
国家自然科学基金资助
No.30170927
No.30070210~~