新一代阳离子聚合物转染试剂(梭华-Sofast)转染效果研究

Study on the Transfection Efficiency of the New Cationic Polymer Transfection Reagent: Sofast

采用DNA延滞实验研究新一代阳离子聚合物梭华 Sofast转染试剂与DNA的结合能力,以不同报告基因(绿色荧光蛋白基因、β 半乳糖苷酶基因和荧光素酶基因),分析比较梭华 Sofast与阳离子脂质体Lipofectamine2000和阳离子聚合物JetPEI、Superfect转染HEK293细胞转染效率和细胞毒性,研究梭华 Sofast转染试剂转染效率.实验结果表明梭华 Sofast与DNA有很强的结合能力;以绿色荧光蛋白基因为报告基因,梭华 Sofast转染率最高,约60%左右,而且在单个细胞中表达的GFP量多;转染细胞荧光素酶活性检测结果表明,当梭华 Sofast与DNA质量比为16时,转染率最高,高于109RLU/mg蛋白,分别是JetPEI,Superfect和Lipofectamine2000转染细胞最高活性的1.6,2.3和5倍;梭华 Sofast转染β 半乳糖苷酶的染色结果显示95%以上的阳性细胞转染率;比较基因转染效率最高时的细胞毒性,在最佳转染剂量时,几种试剂的细胞存活率均在83%以上,而梭华 Sofast为94.23%.在目前市场上销售的转染试剂对HUV EC细胞的转染效果均不理想情况下,梭华 Sofast仍具一定的转染效率,阳性细胞约占20%.研究结果表明梭华 Sofast是一种高转染效率,低细胞毒性的转染试剂.

To study the transfection efficiency of the new cationic transfection reagent: Sofast. DNA arrearage assay was firstly applied to determine the capacity of combination between transfection reagent Sofast and DNA. Then several different report genes (GFP gene , β-galactosidase gene and luciferase gene) were adopted to compare the transfection efficiency to HEK293 cells of Sofast and the toxicity with those of cationic liposome Lipofectamine 2000, cationic polymer Jet PEI and Superfect . In our study, DNA arrearage assay showed the strong combination between Sofast and DNA; Using GFP as report gene, we found that the highest transfection efficiency can be acquired from Sofast among the four transfection reagents, it is about 60%, moreover GFP expressed largely in single cell; Activiation luciferase detection suggested that when the weight ratio of Sofast to DNA was about 16, its transfection efficiency got over 10~9RLU/mg and this was higher than that of the other three. Furthermore, this efficiency is 1.6, 2.3 and 5 folds of the highest efficiency of Jet PEI, Superfect and Lipofetamine 2000, respectively. According to β-galactosidase staining, we found that the positive cell transfection ratio of Sofast got over 95%. On the other hand, the cytotoxicity assay showed that over 83% cell livability could be achieved by Lipofectamine 2000, Jet PEI and Superfect , while 94.23% cell livability by Sofast. Specially, to HUV-EC cell line, Sofast could still maintain about 20% transfection efficiency, though that of all the other reagents available in market now was low.Sofast is a high efficiency but low cell toxicity transfection reagent.