摘要
目的 :以增强型绿色荧光蛋白 (enhancedgreenflu orescentprotein ,EGFP)为报告基因 ,研究lipofectamine 2 0 0 0、Fu GENE6和逆转录病毒载体介导的基因导入系统对视网膜色素上皮细胞的基因转染效率 ,为视网膜及视网膜相关疾病的基因治疗奠定基础。方法 :在体外分离培养人视网膜色素上皮细胞 ,构建含有EGFP基因的表达载体 ,分别应用阳离子脂质体lipofectamine 2 0 0 0、FuGENE6和逆转录病毒三种方法将EGFP基因转染到人视网膜色素上皮细胞中 ,转染后于 4 8h在荧光显微镜下观察EGFP的表达情况。结果 :三种方法介导EGFP基因在人视网膜色素上皮细胞中表达的阳性比例平均分别为 5 2 %、10 %和 72 %。结论 :在体外可通过逆转录病毒系统或阳离子脂质体lipofectamine 2 0 0 0有效地将外源基因导入到人视网膜色素上皮细胞中 ,其中以逆转录病毒系统转染效率为最高。
Objective:Enhanced green fluorescent protein was used as report gene to study the efficiency of gene transfection of lipofectamine 2000,FuGENE6 and retroviral system,which will lay the foundation of gene therapy for retinal and retinal related disease.Methods:To separate and culture human pigment epithelial cells in vitro,construct expression vector containing EGFP,transfect EGFP into human pigment epithelial cells using cationic liposome lipofectamine 2000,FuGENE6 and retrovirus respectively,observe the expression of EGFP under fluorescent microscope in some time.Results:The positive ratio of EGFP expression in human pigment epithelial cells with three methods was 52%,10%,72% respectively.Conclusion:Gene can be delivered into human pigment epithelial cells effectively by retroviral system or cationic liposome lipofectamine 2000,of which transfection efficiency of retroviral system is the most effective.
出处
《眼视光学杂志》
2003年第4期221-223,共3页
Chinese Journal of Optometry & Ophthalmology
基金
浙江省自然科学基金 (ZB0 2 0 2
3 0 2 0 95 )
浙江省科技厅重点课题 ( 2 0 0 3C2 3 0 0 5 )
