电穿孔法介导转染老年人成纤维细胞参数的优化  

Investigation on optimal parameters of electroporation-mediated transfection in elderly fibroblasts

作　　者：王雅珍[1] 杨舟鑫[1] 徐伟红[1] 吕远栋[1] 毛根祥[1] 

机构地区：[1]浙江医院浙江省老年医学重点实验室浙江省老年医学研究所,浙江杭州310013

出　　处：《中国卫生检验杂志》2015年第23期3985-3987,共3页Chinese Journal of Health Laboratory Technology

基　　金：国家自然科学基金(31201040);浙江省医药卫生平台计划项目(2015DTA001)

摘　　要：目的探究采用电穿孔法将质粒pGenesil-1(简称p G)转染老年人成纤维细胞,从中寻找最佳的参数。方法采用电穿孔法将能表达G418抗性蛋白和增强型绿色荧光蛋白(EGFP)的质粒p G转染老年人成纤维细胞。荧光显微镜下观察EGFP表达并测定转染效率、细胞存活率;通过G418筛选得到稳定转染的细胞。结果本实验室电穿孔法介导的老年人成纤维细胞转染的最佳电转条件:电压:240 V;电容:750μF;DNA剂量:8μg。48 h后电穿孔法的转染率接近30%。结论用电穿孔法将质粒pG稳定转染老年人成纤维细胞耗时短,有利于在体外对老年人成纤维细胞进行后续实验。Objective To transfect plasmid p Genesil-1（ p G） into elderly so as to fibroblasts find the best parameters. Methods Electroporation was used for transfect plasmid p G,a plasmid that expresses G418 resistance protein and enhanced green fuluorescent protein（ EGFP）,into elderly fibroblasts. The expression of EGFP was observed and the transfection efficiency was measured after the transfection by fluorescence microscope. The stable transfected cells were screened by G418. Results The optimum conditions for laboratory electroporation-mediated elderly fibroblasts transfection were： power transfer voltage of240 V,capacitance of 750 μF,DNA dose of 8 μg. Transfection efficiency of electroporation after 48 h was about 30%. Conclusion Stably transfecting plasmid p G into elderly fibroblasts with electroporation need short time,and this will help to take advantage of the elderly fibroblasts for subsequent experiments.

关 键 词：老年人 成纤维细胞 电穿孔 

分 类 号：R329.3[医药卫生—人体解剖和组织胚胎学]