CUY21EDITⅡ超级多脉冲活体/细胞电转化仪转染外源基因至神经干细胞的优化方案  

Optimization protocol for transfecting exogenous gene into neural stem cells by CUY21EDITⅡsuper multi pulse in vivo / cell electroporator

作　　者：王经纬[1,2] 田景瑞[1] 赵彬[1] 田艳霞[1] 高俊玲[1] 伊红丽[2] 江小华[1] 

机构地区：[1]河北联合大学基础医学院 [2]河北联合大学附属医院,唐山063000

出　　处：《神经解剖学杂志》2015年第3期322-326,共5页Chinese Journal of Neuroanatomy

基　　金：河北省自然基金(H2012209035);唐山市科技研究项目(111302111b)

摘　　要：目的:优化CUY21EDITⅡ电转仪介导真核表达载体p EGFP-N1转染大鼠来源的神经干细胞(NSCs)的转染条件,提高外源基因转染NSCs的转染效率,为后续试验提供基础。方法:在保持驱动电压(Pd V)、脉冲时间、循环次数等条件不变的情况下,通过改变电转电压(Pp V),将已构建好的表达绿色荧光蛋白的重组载体p EGFP-N1以CUY21EDITII电转仪转染导入NSCs,转染后48 h在荧光显微镜下观察荧光并计算转染效率。结果:p EGFP-N1质粒在Pp V为350 V,Pd V为20 V,脉冲时间为10 ms,循环次数为20次的转染条件下,可获得较理想的转染效率,转染效率平均为81.0%,细胞存活率为72.0%。结论:通过优化转染条件,CUY21EDITⅡ电转仪能提高p EGFP-N1转染NSCs的转染效率,与脂质体相比优势明显。Objective： By optimizing the conditions of transfecting eukaryotic expression vector p EGFP-N1 into rat neural stem cells（ NSCs） with CUY21 EDITII electroporator,to elevate the transfection efficiency and provide a basis for the follow-up experiments. Methods： Keeping Pd V,pulse time and cycle time etc. unchanged,just changing Pp V,p EGFPN1 plasmid which expresses green fluorescence protein（ GFP） was transfected into NSCs by CUY21 EDITII electroporator.Then the transfection efficiency was calculated through counting the NSCs with green fluorescence under the fluorescence microscope at 48 h after transfection. Results： Under the condition of 350 V Pp V,20 V Pd V,10 ms pulse time and 20 cycles,the average transfection rate was 81. 0% and the survival rate was 72. 0%. The transfection efficiency was satisfactory. Conclusion： The transfection rate of p EGFP-N1 into NSCs using CUY21EDITⅡ electroporator could be elevated by optimizing the transfection conditions,and was higher than using lipofectamine.

关 键 词：神经干细胞 电穿孔 脂质体 转染 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]