三种转染试剂对PEF细胞与BHK细胞转染效率的比较

Transfection Efficiency of Three Transfection Reagents to PEF Cells and BHK Cells

【目的】比较三种转染试剂对原代细胞和传代细胞的转染效率,选择合适的转染试剂。【方法】以EGFP为报告基因,分别用脂质体、磷酸钙、罗氏试剂转染PEF细胞和BHK细胞。通过改变转染试剂和质粒DNA的剂量,利用流式细胞仪分析转染效率,从而确定最佳的转染试剂。【结果】对于PEF细胞,脂质体和磷酸钙转染效果不理想,细胞死亡率高,绿色荧光表达量少。罗氏转染试剂相对较好,细胞死亡率低,并且保持原有细胞形态。对于传代细胞BHK细胞而言,各转染试剂均可达到理想转染效果。【结论】不同的细胞对转染试剂的选择也不同,传代细胞BHK用脂质体即可达到理想的效果。利用罗氏试剂可以提高转染效率,但不明显。而对于原代细胞PEF,脂质体与磷酸钙试剂的转染效率很低。相比较而言,罗氏转染试剂效果更好些,细胞毒性小,转染率在10%～12%。

[ Objective] The project aims to compare the transfection efficiency of the three transfection reagents to primary cells and passage cells and choose the appropriate transfection reagents. [ Method] EGFP as a reporter gene, the liposome, calcium phosphate and the Roche reagent transfection were transfected into PEF cells and BHK cells respectively. The optimal transfection reagent was determined by analyzing the dose of transfection reagent and plasmid DNA with flow cytometer. [ Result] For PEF ceils, transfectamine 2000 and calcium phosphate transfection effect was not ideal with cell high mortality and the less expression of green fluorescent. While the transfection effect of Roche transfection reagent was relatively better, in which cell mortality was low, and cell morphology was normal. For the passage BHK cells, various transfection reagents might reach the ideal transfection effect. [ Conclusion ] The selection of the transfection reagent was different for different ceils and the passage BHK ceils could achieve the desired transfection effect with transfectamine 2000. Roche reagents could improve the efficiency of transfection, but not obviously. For primary PEF cells, cytotoxicity of transfectamine 2000 was greater, and difficult to transfect. While the transfection effect of Roche reagent was better, with low cytotoxicity. The transfection rate was between 10% -12%.