中文摘要：

目的了解不同大小质粒和不同基因转染法对人KC基因导入效率的影响。方法采用脂质体转染法、阳离子多聚物转染法、电穿孔联合细胞核转染试剂转染法、慢病毒感染法，分别将不同大小的质粒[pSUPER-增强型绿色荧光蛋白（EGFP）、pEGFP—N2、pHSER-绿色荧光蛋白（GFP）、ploxP-EGFP]导入人永生化KC株HaCaT细胞和人胚肾细胞株293FT细胞（后者为对照）。于倒置荧光显微镜下观察GFP的表达，计算转染率。结果（1）采用脂质体转染法可将4种质粒导入HaCaT细胞（转染率1．0％～3．3％）及293FT细胞（转染率80．0％～84．7％）。（2）阳离子多聚物转染法亦可将4种质粒导入HaCaT细胞（转染率为1．0％～3．7％）和293FT细胞（转染率81．3％～86．7％）。（3）采用电穿孔联合细胞核转染试剂转染法，可以将2种较小片段质粒pSUPER—EGFP和pEGFP-N2导入HaCaT细胞，转染率分别为22．3％和19．0％；而2种较大片段质粒pHSER-GFP和ploxP-EGFP的转染率分别为4．0％和3．3％。（4）pHSER-GFP经慢病毒包装后，导入HaCaT细胞的转染率高达97．0％，明显优于前3种转染法。结论脂质体转染法、阳离子多聚物转染法较难将外源性基因导入人KC，慢病毒感染法的转染率明显优于电穿孔联合细胞核转染试剂转染法；不同大小质粒对转染率有明显影响

英文摘要：

Objective To observe the effect of plasmids in different size and gene transfection protocol on efficiency of introducing gene into human KC. Methods Four plasmids in different size, including pSUPER-enhanced green fluorescent protein （EGFP） , pEGFP-N2, pHSER-green fluorescent protein （GFP） and ploxP-EGFP, were transfected into immortal human KC line （HaCaT） and human embryo kidney cell line （293FT） separately following transfection protocols of liposome （LTP）, cation polymerizer （CPTP） , electroporation combined with nucleus transfection agent （ETP） and lentivirus. 293FT was used as control. GFP expression was observed under inverted fluorescence microscope. The transfection efficiency （TE） was calculated. Results （1） The four plasmids could be introduced into HaCaT （TE, 1.0%- 3.3% ） and 293FT （ TE, 80.0% -84.7% ） following LTP. （2） The four plasmids could also be introduced into HaCaT （TE, 1.0% -3.7%） and293FT （TE, 81.3% -86.7%） followingCPTP. （3） Two shorterplasraids （ pSUPER-EGFP and pEGFP-N2） could be introduced into HaCaT by ETP with higher TE than the other two longer plasmids （ pHSER-GFP and ploxP-EGFP） , which were 22.3% and 19.0% vs. 4.0% and 3.3% , respectively. （4） pHSER-GFP packaged by lentivirus could be introduced into HaCaT with the TE reaching 97.0% , which surpassed the above three protocols. Conclusions It is difficult to introduce exogenous gene into human KC by LTP or CPTP; TE of lentivirus transfection protocol apparently surpasses ETP. Plasmid size can greatly affect TE.