腺相关病毒介导增强型绿色荧光蛋白基因体外转染兔结膜上皮细胞  

Transfection of rabbit conjunctival epithelial cells with adeno-associated virus mediated enhanced green fluorescent protein

作　　者：苏莉[1] 崔红平[2] 李艳红[2] 高新蕊[2] 

机构地区：[1]上海交通大学附属第一人民医院眼科,上海200080 [2]同济大学附属上海市第十人民医院眼科,上海200072

出　　处：《第二军医大学学报》2008年第12期1499-1502,共4页Academic Journal of Second Military Medical University

基　　金：国家自然科学基金(30570954)~~

摘　　要：目的:研究2型重组腺相关病毒(recombinant adeno-associated virus 2,rAAV2)载体介导增强型绿色荧光蛋白基因(enhanced green fluorescent protein,EGFP)对体外培养兔结膜上皮细胞的转染和表达情况,为后续研究提供依据。方法:体外培养兔结膜上皮细胞,rAAV2-EGFP按感染复数(multiplicity of infection,MOI)为104、105、106转染第2代兔结膜上皮细胞,转染后第1、3、5、7日倒置荧光显微镜下观察细胞中EGFP表达情况,计算转染率。MTT方法检测rAAV2-EGFP转染对细胞增殖的影响。结果:随着MOI值增大及转染时间延长,EGFP表达效率逐渐增高,转染后第7～8日达到高峰并维持。MTT检测各MOI组与对照组差别无统计学意义。结论:rAAV2载体可以介导EGFP基因高效稳定转染兔结膜上皮细胞,并且对细胞增殖无影响。Objective： To transfect rabbit conjunctival epithelial cells in vitro with recombinant adeno-associated virus 2- mediated enhanced green fluorescent protein （rAAV2-EGFP）, so as to lay a foundation for studying the proliferation and differentiation of conjunctival epithelial stem cells. Methods： Rabbit conjunctival epithelial cells were cultured in vitro. The second passage of rabbit conjunctival epithelial cells were transfected with rAAV2-EGFP in various multiplicity of infection （MOI：104 ,105 ,10G ）. The expression of green fluorescent protein was examined 1,3,5, and 7 days after transfection and the transfection rate was calculated. MTT assay was used to study the influence of rAAV2-EGFP on the growth of cells. Results： With the increase of MOI value and the prolongation of transfection, the expression of EGFP was gradually increased and reached its peak on 7-8 days after transfection. MTT assay showed that there was no significant difference between the MOI group and the control group. Conclusion： rAAV2-EGFP can efficiently transfect rabbit conjunctival epithelial cells and have no influence on the proliferation of the cells.

关 键 词：腺相关病毒 绿色荧光蛋白 结膜上皮细胞 荧光蛋白质示踪 

分 类 号：R943.43[医药卫生—药剂学]