阳离子脂质体介导基因转染的实验教学改革

Experiment Teaching Reform of Cationic Liposome-mediated Gene Transfection

通过优化细胞转染条件,探讨脂质体与DNA的复合比例、复合时间以及转染时间对脂质体介导的基因转染效率的影响。实验采用凝胶阻滞电泳分析脂质体完全包裹DNA时二者的复合比,通过转染绿色荧光蛋白报告基因(pEGFP)研究脂质体/pEGFP的复合比例、复合时间对细胞转染效率的影响。激光扫描共聚焦成像进一步分析转染时间对细胞摄取DNA量的影响。实验结果表明,脂质体和DNA的复合比例为1.25时,细胞的转染效率最高。转染4 h后,大量DNA被细胞摄取,此时可以更换新鲜的培养基。

By optimizing the conditions of gene transfection, we explored the effect of the gene transfection conditions, such as [ liposome ] / [ DNA ] ratios, incubation time and exposure time of the complexes with cells, on gene transfection efficiency. Gel retardation assay was used to fix the [ liposome] / [ DNA] ratios, and then the effects of such combination ratios and times on the gene transfection efficiency were examined. Moreover, the quantity of DNA uptaked by cellular was further analyzed by using the confocal laser scanning microscope. According to the results, when the ratio of [ liposome] / [ pEGFP] is 1.25, gene transfection efficiency is the highest. After 4 hours of transfection, most of DNAs have been uptaked by the cells. It＇s time to replace fresh culture medium.