摘要
目的研究用慢病毒载体进行基因转染对小鼠胚胎干细胞(ESC)特性的影响。方法慢病毒载体经293FT细胞包装后转染小鼠ESC,荧光显微镜下挑选和观察所得阳性克隆,检测ESC特异性标志物Oct-4、SSEA-1的表达,通过拟胚体(EB)诱导体外分化,RT-PCR检测三胚层的分化。结果形态学观察可见所得绿色荧光蛋白(GFP)阳性细胞呈克隆样鸟巢状生长,免疫组化可见GFPESC表达ESC特异性标志Oct-4、SSEA-1,能形成EB,RT-PCR检测到甲胎蛋白(AFP)、巢蛋白(Nestin)和血管内皮生长因子受体(Flk-1)的表达。结论慢病毒载体介导的基因转染不影响小鼠ESC的自我更新特性及多向分化潜能。
Objective To study the influence of gene transfection by lentiviral vector on mice embryonic stem cell （ESC） features. Methods Transfection of mice ESC was made by lentiviral vector after 293FT cell packing. Positive clone was chosen and observed under fluorescence microscope. Expressions of Oct-4 and SSEA-1 as ESC specific markers were detected. In vitro differentiation was induced by embryoid body （EB）, and RT-PCR was applied to detect differentiation of triploblastica. Results Green fluorescent protein （GFP） positive cell showed colon bird-nest growth by morphological observation. Immunohistochemistry showed Oct-4 and SSEA-1 as ESC specific markers could form EB. Expressions of alpha fetoprotein （AFP）, nestin and vascular endothelial growth factor receptor （Flk-1） could be detected by RT-PCR. Conclusion Gene transfection mediated by lentiviral vector has no influence on self-renewal feature and multi-directional differentiation potency of mice ESC.
出处
《中国实用医药》
2015年第11期3-5,共3页
China Practical Medicine
关键词
胚胎干细胞
慢病毒载体
基因转染
绿色荧光蛋白
Embryonic stem cell
Lentiviral vector
Gene transfection
Green fluorescent protein
