聚乙烯亚胺介导三质粒共转染HEK293细胞的条件优化  

Optimization of Triple Plasmids Transfection into HEK293 Cells Mediated by Polyethylenimine

作　　者：付强[1,2] 李艳[3] 郑昭芬[1,2] 刘爱忠[3] 袁振华[4] 彭建强[1,2] 何晋[1,2] 

机构地区：[1]湖南师范大学第一附属医院 [2]湖南省人民医院心血管内科,长沙410005 [3]中南大学湘雅公共卫生学院流行病学与卫生统计学系,长沙410078 [4]中国科学院深圳先进技术研究院生物医药与技术研究所,深圳518055

出　　处：《生物医学工程学杂志》2015年第1期137-141,共5页Journal of Biomedical Engineering

基　　金：长沙市民生科技支撑专项基金资助项目(K1106022-31)

摘　　要：本研究用聚乙烯亚胺（PEI）介导表达绿色荧光蛋白（GFP）的rAAV包装系统（pAAV-CMV-GFP、pAAVRC和pHelper）转染人胚肾293细胞（HEK293细胞）。用ImagePro Plus6.0图像分析软件计算表达GFP的细胞比例来测定1、2、3、4、5、6μg/孔不同质粒剂量,1∶1、3∶1、5∶1、7∶1不同PEI/质粒比值及转染12、24、36、48、60、72h不同时间三个变量对转染效率的影响,以优化PEI介导三质粒共转染HEK293细胞的条件。结果显示,质粒剂量为4μg/孔、PEI/质粒比值为3∶1-5∶1时转染效率维持较高水平;转染效率-转染时间曲线为S型增长曲线,转染效率于60h达到平台期。优化PEI转染条件能增加三质粒共转染HEK293细胞的转染效率。In the present study,packaging system composed of pAAV-CMV-GFP,pAAV-RC and pHelper were transfected into human embryonic kidney 293cells（HEK293cells）mediated by polyethyleneimine（PEI）to explore an optimal transfection condition.Different total plasmid DNA dosages（1,2,3,4,5,6μg）and different PEI/Plasmid ratios（1∶1,3∶1,5∶1,7∶1）were tested with detection of green fluorescence protein（GFP）with ImagePro Plus6.0Software.Then transfection efficiency of the optimized transfection system was further observed for different time periods（12,24,36,48,60,72h）.The results showed that total plasmid dosage of 4μg/well with PEI/plasmid ratio of 3∶1-5∶1 was an efficient transfection condition.Transfection efficiency-time curve was an Sshaped curve.Transfection efficiency reached a plateau at 60 hafter transfection.The optimized conditions for PEImediated transfection at the optimal time result in enhanced transfection efficiency of triple plasmid into HEK293 cells.

关 键 词：聚乙烯亚胺 腺相关病毒 HEK293细胞 转染效率 条件优化 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]