Lipofectamine~ LTX转染Jurkat细胞条件的优化  

Optimization of Condition for Transfection of Jurkat Cells with Lipofectamine~ LTX

作　　者：牛超[1] 田代印[2] 符州[2] 王莉佳[1] 张明香[2] 

机构地区：[1]重庆医科大学儿童发育疾病研究省部共建教育部重点实验室,重庆400014 [2]重庆医科大学附属儿童医院呼吸科,重庆400014

出　　处：《中国生物制品学杂志》2011年第12期1507-1510,1517,共5页Chinese Journal of Biologicals

基　　金：国家自然科学基金(30672268)

摘　　要：目的优化阳离子脂质体Lipofectamine誖LTX转染悬浮细胞Jurkat的条件。方法将带有绿色荧光蛋白(GFP)基因的两种质粒pIRES2-EGFP和pIRES2-EGFP-STAT6作为基因载体,采用Lipofectamine誖LTX转染Jurkat细胞,绘制细胞生长曲线,检测培养基新鲜度对细胞生长的影响及细胞培养时间、脂质体与质粒DNA的比例、细胞接种密度、不同量培养基等对转染效果的影响,比较瞬时转染效果。结果 Jurkat细胞生长速度与细胞密度密切相关,转染前1 d换液,可保证细胞的良好生长状态,培养基新鲜度对细胞生长影响不大。脂质体体积与质粒质量之比为2.75μl∶500 ng时,其转染效果明显优于其他各组。适当加大转染时Jurkat细胞的铺板密度,可以明显提高转染试剂的利用率。转染48 h后,300μl培养基/孔组的绿色荧光阳性率为600μl培养基/孔组的(1.4±0.2)倍,差异有统计学意义(P<0.05)。转染后24 h,Lipofectamine誖LTX体积与质粒质量各比例组转染效果差异无统计学意义(P>0.05);转染48 h后,各比例组转染效果差异有统计学意义(P<0.05),且质粒pIRES2-EGFP的转染效果明显优于pIRES2-EGFP-STAT(6P<0.05)。结论优化了Lipofectamine誖LTX转染悬浮细胞Jurkat的条件,为悬浮细胞的转染提供了参考。Objective To optimize the condition for transfection of Jurkat cells with Lipofectamine LTX.Methods Jurkat cells were transfected with Lipofectamine LTX using plasmids pIRES2-EGFP and pIRES2-EGFP-STAT6,both carrying green fluorescent protein（GFP） gene,as vectors,based on which the cell growth curve was plotted,the effect of freshness of medium on cell growth as well as effects of time for culture,ratio of Lipofectamine LTX to plasmid DNA,density of cells inoculated and volume of medium on transfection efficacy were evaluated,and the transient transfection efficacies were compared.Results The growth rate of Jurkat cells was closely related to the cell density.The change of medium 1 d before transfection ensured the normal growth state of cells.However,the freshness of medium showed no significant effect on cell growth.When the ratio of Lipofectamine LTX volume to plasmid mass was 2.75 μl ∶ 500 ng,the transfection efficacy was satisfactory.Properly increased density of Jurkat cells increased the utilization rate of transfection reagent.Forty-eight hours after transfection,the transfection efficacy of cells inoculated in 300 μl of medium onto each well of culture plate was（1.4 ± 0.2） times of that in 600 μl of medium,which showed significant difference（P 0.05）.The transfection efficacies at various ratios of Lipofectamine LTX volume to plasmid mass showed no significant difference 24 h（P 0.05） while showed significant difference 48 h（P 0.05） after transfection.The transfection efficacy of plasmid pIRES2-EGFP was significantly superior to that of plasmid pIRES2-EGFP-STAT6（P 0.05）.Conclusion The condition for transfection of Jurkat cells with Lipofectamine LTX was optimized,which provided a reference for the transfection of suspension cells.

关 键 词：阳离子脂质体 悬浮细胞 JURKAT细胞 转染 

分 类 号：Q78[生物学—分子生物学]