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目的优化LipofectamineTM2000介导外源基因转染Neuro-2a(N-2a)细胞的条件,以提高转染效率。方法将携带增强型绿色荧光蛋白(Enhanced green fluorescent protein,EGFP)的真核表达载体pEGFP-N1在LipofectamineTM2000介导下转染N-2a细胞,对转染时间、细胞融合度、质粒质量与脂质体体积比例及培养基pH值进行优化,荧光显微镜观察EGFP的表达,并计算转染效率。结果在细胞融合度达80%,质粒质量与脂质体体积比为1∶3的条件下转染48 h,EGFP的阳性率最高(P<0.05);培养基pH值为8.0左右时,EGFP的阳性率较高,且细胞凋亡率较低。结论优化了LipofectamineTM2000转染N-2a细胞的条件,提高了转染效率,为外源基因高效转染N-2a细胞奠定了基础。
Objective To optimize the conditions of LipofectamineTM2000-mediated transfection of Neuro-2a (N-2a) cells in order to improve transfection efficiency. Methods The eukaryotic expression vector pEGFP-N1 carrying Enhanced Green Fluorescent Protein (EGFP) was transfected into N-2a cells with LipofectamineTM2000. The transfection time, cell fusion, plasmid quality and lipid Volume ratio and medium pH value were optimized, the expression of EGFP was observed by fluorescence microscopy, and the transfection efficiency was calculated. Results The positive rate of EGFP was the highest (P <0.05) when the degree of cell fusion was 80% and the mass ratio of plasmid to liposome was 1: 3 for 48 h. When the pH was about 8.0, EGFP The positive rate is higher, and the apoptosis rate is lower. Conclusion The conditions of LipofectamineTM2000 transfection of N-2a cells were optimized and the transfection efficiency was improved, laying a foundation for efficient transfection of N-2a cells with exogenous gene.