电穿孔转染法与脂质体转染法对K562和HepG2细胞转染效率的影响  

Effect of lipofectin and electroporation transfection method on cells transfection efficiency of K562 and HepG2 cells

作　　者：隋娟[1] 张高丽[1] 李平法[1] 于海川[1] 

机构地区：[1]新乡医学院医学检验学院,河南新乡453003

出　　处：《新乡医学院学报》2015年第8期698-701,共4页Journal of Xinxiang Medical University

基　　金：国家自然科学基金项目(编号:31301135);河南省教育厅科学技术研究重点项目(编号:14B320015)

摘　　要：目的通过对电穿孔转染法和脂质体转染法在K562、HepG2细胞中的转染效率比较,探索最佳的细胞转染方法和条件。方法以K562、Hep G2细胞为研究对象,采用电穿孔转染法和脂质体转染法分别转染带有绿色荧光的质粒PEGFP-N1和无荧光的质粒p CDNA3.1、p CDNA3.1-EOS基因,荧光显微镜下观察PEGFP-N1转染效果,计算转染效率;收集各培养组细胞,裂解后提取蛋白,采用Western blot方法检测基因转染后的蛋白表达。结果当脉冲20 ms、电阻90Ω、电压150 V时进行电穿孔,转染质粒PEGFP-N1,K562细胞可以得到较高的转染率;当脉冲20 ms、电阻90Ω、电压250 V时,转染质粒PEGFP-N1,Hep G2细胞可以得到较高的转染效率。在K562细胞和Hep G2细胞中,电穿孔转染法的转染效率均显著高于脂质体转染法(P<0.05)。电穿孔转染法转染PEGFP-N1后K562细胞和Hep G2细胞的存活率显著低于脂质体法(P<0.05)。Western blot结果显示电穿孔转染法蛋白表达量高于脂质体转染法(P<0.05)。结论最佳条件下电穿孔转染法是一种高效的基因转染方法,对比较难转染的悬浮细胞效果更佳。Objective To explore the optimum cell transfection method and condition by comparing the transfection ef- ficiencies of lipofectin and electroporation transfeetion method in human K562 and HepG2 cells. Methods The K562 and HepG2 ceils were selected as research object. The electroporation transfection method and lipofectin transfection method were used to transfect the plasmid pEGFP-N1 （with green fluorescent） and plasmid pcDNA3. 1, pCDNA3. 1-EOS gene（ without green fluorescence）respectively. The transfection efficiency of pEGFP-N1 was observed by fluorescence microscope, and then calculated the transfection efficiency. The cells in each cultured group were collected and splitter, then extracted the protein; Western blot was used to detect protein expression after gene transfection. Results The K562 cells could get high transfection efficiency under a pulse of 20 ms ,90 Ω resistance ,voltage 150 V conditions for transfecting plasmid PEGFP-N1 by electropora- tion;the HepG2 cells could get high transfection efficiency under a pulse of 20 ms ,90 Ω resistance, voltage 250 V conditions for transfecting plasmid PEGFP-N1 by electroporation. The transfection efficiency of electroporation transfection method was significantly higher than the lipofectin transfection method in both K562 cells and HepG2 cells （ P 〈 0.05 ）. The survival rate of K562 cells and HepG2 cells after transfecting PEGFP-N1 by electroporation transfection method was significantly lower than li- pofectin transfection method（P 〈 0.05 ）. Western blot results showed that the electroporation transfection method had higher protein expression than liposome transfection method （ P 〈 0. 05 ）. Conclusion Under the optimum conditions, electroporation is a high-performance method for gene transfection, and it is more effective for the suspension cells.

关 键 词：电穿孔 脂质体转染 K562细胞 HepG2细胞 

分 类 号：Q813.1[生物学—生物工程]