摘要
目的:原代海马神经元是原代细胞中比较难转染的细胞,为了得到比较高的转染效率,通过比较Li-pofectamine2000转染法和大鼠神经元核电转法(rat neuron nucleofector kit)转染效率及转染前后细胞的存活率,来探讨Nucleofector Kit核电转法的高效性。方法:选取胚胎18 d(E18)大鼠的海马神经元进行原代培养,获取海马神经元单细胞悬液后,于种植前和种植24 h后分别采用Nucleofector Kit和Lipofectamine 2000转染红色荧光蛋白(red fluorescence protein,DsRed)质粒。神经元转染48 h后分别采用轴突标记物Tau-1进行免疫荧光染色鉴定神经元。神经元的存活率采用台盼兰进行检测。结果:Lipofectamine2000的基因转染率仅为5.34%,而大鼠神经元Nucleofector Kit的转染率为44.54%,后者为前者的8.34倍。Lipofectamine2000转染细胞前后的存活率分别为98.56%和91.44%,而Nucleofector Kit组分别为98.26%和74.02%。结论:大鼠神经元核电转法能高效稳定的转染原代海马神经元。
Objective： It is very difficult for primary hippocampal neurons to be transfected.In order to get high transfection efficiency,we transefected the neurons with two methods,which are lipofectamine2000 and rat neuron nucleofector kit.We compared the cellular transfected efficiency and cellular survival rate.Methods： At embryo 18 days（E18） rats were used to culture primary hippoampal neurons.Before and after neuronal plating for 24 h,nucleofector kit and lipofectamine2000 were transfected into the neurons with red fluorescence protein（DsRed） plasmid,respectively.After transfection for 48 h,axon maker-Tau-1 was immunofluorescencely stained to evaluate the neurons,and the neuronal survival rate was detected with trypan blue staining.Results： The efficiency rate of lipofactamine 2000 was only 5.34%,however,it was up to 44.54% with nucleofector kit,which was 8.34 times than the former.The survival rate before and after transfection was 98.56% and 91.44%,whereas they was 98.26% and 74.02% respectively in nucleofector transfected group.Conclusion： The method of using the rat neuron nucleofector kit can transfected high efficiency for primary hippocampal neurons.
出处
《神经解剖学杂志》
CAS
CSCD
北大核心
2011年第6期689-692,共4页
Chinese Journal of Neuroanatomy
