Optimization of N2a cell transfection mediated by liposome / 中国组织工程研究

ABSTRACT

BACKGROUND:Cationic liposome-mediated celltransfection is reliable and repeatable. However the transfection efficiency is often low. OBJECTIVE:To study the optimized methods for gene transfection mediated by liposome into N2a cells (mouse neuroblastma cells). METHODS:Using traditional adherent method and improved suspension method, 500 ng recombinant plasmid pcDNA3-GFP carrying green fluorescence protein was transfected into N2a cells in 24-wel culture plate, which was mediated by 1.5μL Lipofectamine?LTX Reagent. The expression of green fluorescent protein was observed by inverted fluorescence microscope, and the transfection efficiencies at different transfection ways were calculated. By using improved suspension transfection method, 500 ng plasmid DNA was transfected with different doses of Lipofectamine?LTX Reagent (1.0, 1.5, 2.0, 2.5μL). The optimal ratio of liposome and DNA was explored. RESULTS AND CONCLUSION:The transfection efficiency of suspension transfection method was significantly higher than that of the tranditional adherent method (P<0.01) when using 1.5μL liposome/500 ng DNA. The transfection efficiency of the 1.0, 1.5, 2.0, 2.5μL Lipofectamine?LTX on 500 ng plasmid DNA was respectively (76.60±3.85)%, (80.00±4.17)%, (88.00±5.89)%, (54.96±4.23)%. It showed the 500 ng DNA and 2.0μL liposome achieve the highest transfection efficiency.