增强型绿色荧光蛋白基因在人胚胎视网膜前体细胞中的表达  

The Expression of Enhanced Green Fluorescent Protein Gene on Cultured Human Fetal Retinal Progenitor Cells

作　　者：康前雁[1] 刘勇[1] 张瑾[1] 刘建新[1] 石秦东[1] 田英芳[1] 肖新莉[1] 张殿增[1] 钱亦华[1] 

机构地区：[1]西安交通大学医学院环境与疾病相关基因教育部重点实验室神经科学研究中心

出　　处：《眼科学报》2006年第1期54-58,共5页Eye Science

基　　金：国家自然科学基金资助项目(30170300;30200291);陕西省科技攻关资助项目(2002K10-G8);西安交通大学自然科学基金资助项目

摘　　要：目的:研究增强型绿色荧光蛋白基因(enhanced green fluorescent protein,EGFP)转染人胚胎视网膜前体细胞的转染效率及瞬时表达情况,建立人胚胎视网膜前体细胞的示踪方法,为视网膜前体细胞移植提供示踪依据。方法:在大肠杆菌中扩增pEGFP-N1质粒,通过磷酸钙介导法转染人胚胎视网膜前体细胞．应用流式细胞仪检测其转染效率,荧光显微镜和激光共聚焦显微镜观察其瞬时表达情况。结果:pEGFP-N1在基因转染24 h细胞转染率为28．98％,48 h为32．45％,72 h为30．40％,对照未转染细胞24 h、48 h分别为2．50％和2．04％:体外观察培养细胞转染24 h已有绿荧光表达,48-72 h细胞荧光强度最强,96 h荧光强度减弱。结论:pEGFP-N1质粒能有效转染人胚胎视网膜前体细胞,其转染效率可达到30％,是人胚胎视网膜前体细胞较为理想的瞬时表达载体和细胞示踪报告分子。Purpose ：To investigate the transfer efficiency and transient expression of plasmid vector coding enhanced green fluorescent protein （EGFP）gene pEGFP-N1 which was transferred into primary cultured human fetal retinal progenitor cells. To build up a tracking method for study of retinal progenitor cells transplantation. Methods：pEGFP-N1 plasmid was amplified in E.coli, primary cultured human fetal retinal progenitor cells transferred with pEGFP-N1 by means of CaC12.The transfer efficiency was evaluated by flow cytometry. Transient expression in vitro was observed by fluorescent microscope and laser scanning confocal microscope. Results：The transfer efficiency of pEGFP-N1 into primary cultured human fetal retinal progenitor cells could maximumly reach to 32.45% during 48 hours. There was obvious expression of pEGFP-Nlin vitro at 48th hour after transferring. Conclusion ：pEGFP-N1 is an ideal transient expression vector for primary cultured human fetal retinal progenitor cells in vitro. It is a useful tracer for transplantation of retinal progenitor cells.

关 键 词：绿色荧光蛋白 视网膜前体细胞 基因转染 胚胎 基因表达 

分 类 号：R774.6[医药卫生—眼科]