影响非洲绿猴肾细胞脂质体转染效率的因素

The critical parameters affecting Liposome-mediated transfection of Vero cells

已有实验表明，细胞转染效率可能决定于ＤＮＡ－脂质体复合物的形成以及所转染的细胞种类。利用脂质体ＬｉｐｏｆｅｃｔＡＭＩＮＥ，研究了影响Ｖｅｒｏ细胞转染效率的参数如ＤＮＡ和脂质体的用量，转染的细胞数量以及细胞暴露于ＤＮＡ－脂质体复合物的时间长度。通过检测报告基因β－半乳糖苷酶（β－ｇａｌ）的表达，发现最高转染率只在一较窄范围内获得。β－ｇａｌ的表达随脂质体量增加而显著增加。在标准转染条件下，增加ＤＮＡ用量和延长细胞暴露时间反而使转染效率下降。转染细胞数量为４×１０４细胞／２４孔时，细胞转染效率最高。

Previous studies have shown that the level of gene expression achieved may be dependent on the formulation of the DNA-liposome complex and the cell type transfected. We have investigated the effect of parameters such as DNA, liposome dose, number of cells transfected and exposure time of the cells to the DNA-liposome complexes on the level of transgene expression in Vero cells using lioposome lipofectAMINE. A narrow range of conditions was founded to produce maximal level of transgene expression, as detected using the reporter molecule β-galactosidase(β-gal). β-gal expression was significantly enhanced as the dose of liposome increased. Under standard transfection conditions, increasing DNA amount and exposure time of the DNA-liposome complexes with cells resulted in decreased transgene expression. Over a wide range of the number of cells transfected, maximal gene expression was found at 4×10 4 cells/24-well. The maximal level of expression that could be produced using fully optimized transfection conditions, however, was still highly dependent on each cell type analyzed.