摘要
本研究采用电转染法将pmaxGFP-gcc质粒通过Lonza电转仪转染入体外培养的水牛精原干细胞。通过将7组电脉冲程序的转染结果进行对比,得出水牛精原干细胞存活率与转染率最高的组别,并确定对应的最佳的电转染程序为EH100,48 h后,流式细胞仪筛选获得此电转染程序下有19.1%的GFP;细胞且转染细胞存活率达到74.2%。本电转染实验操作简便,转染效率高,可重复性好。本实验中的电转染的方法可以用于普通方法难以转染的干细胞,为其他物种的精原干细胞转基因研究提供研究基础和参考。
In this research,pmaxGFP-gcc plasmid was transfected into buffalo spermatogonial stem cells cultured in vitro by Lonza nucleofector.The results showed that the highest survival rate and transfection rate of buffalo spermatogonial stem cells were obtained after comparing the transfection results of seven groups of electrical pulse optimization procedure and the corresponding optimal electrotransfection procedure was determined to be EH100,a rate of 19.1%of GFP+cells were screened by flow cytometry after 48 hours of that electrotransfection and the survival rate of transfected cells reached 74.2%.The procedure of this experiment is simple,high transfection efficiency and good repeatability.This method can be used to transfect stem cells which are difficult to transfect by common methods,and provides the basis and reference for the transgenic research of spermatogonial stem cells from other species.
作者
杨欢
李婷婷
彭腊如
谢龙
杨小淦
陆阳清
梁兴伟
卢晟盛
卢克焕
Yang Huan;Li Tingting;Peng Laru;Xie Long;Yang Xiaogan;Lu Yangqing;Liang Xingwei;Lu Shengsheng;Lu Kehuan(State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources,Guangxi University,Nanning,530004; Guangxi High Education Laboratory for Animal Reproduction and Biotechnology,College of Animal Science&Technology,Guangxi University,Nanning,530004)
出处
《基因组学与应用生物学》
CAS
CSCD
北大核心
2021年第7期2455-2460,共6页
Genomics and Applied Biology
基金
广西优秀博士学位论文培育项目(YCBZ2013008)
广西壮族自治区优秀博士研究生出国研修项目和国家自然科学基金(31460601)
关键词
电转染
水牛精原干细胞
转染效率
Electrotransfection
Buffalo spermatogonial stem cells
Transfection efficiency
