目的 探索以绿色荧光蛋白(GFP)为报告基因的重组质粒pEGFP/SDF-1a/54/KDEL电穿孔悬浮培养的Molt-4细胞的高效转染条件.方法 通过控制脉冲幅度、脉冲电压、细胞生长状态等三个主要条件,采用不同条件组合后用电穿孔法将转入悬浮培养的Molt-4,用荧光显微镜和流式细胞仪观察转染率.结果 传代后第二天的Molt-4细胞在260 V、950 μF得到最大转染率51.2%.荧光显微镜下可观察到被转染目的基因的细胞发出绿色荧光.结论 优选260V、950μF和良好的细胞生长状态等条件下的Molt-4,可获得目的基因SDF-1a/54/KDEL的最高转染效率.%Objective To optimize the electroporation parameters for leukemia cell lines Molt-4 cultured in suspension with reconstruction plasmid pEGFP/SDF-1a/54/KDEL (greenfluorescent protein as a reporter gene). Methods pEGFP/SDF-1a/54/KDEL plasmid was transferred into MOLT-4 cells by electroporation with differently experimental conditions such as pulse amplitude, the voltage and the state of cell vitality. The transfection efficiencies were evaluated by flow cytometry (FCM) and fluorescent microscopy. Results The highest electroporation transfection rate (51.2%) was achieved under the condition of voltage 260 V, 950 μF and the vigorous state of MOLT-4 cells. Conclusion Optimized electroporation parameters 260V, 950 μ F and the vigorous state of MOLT-4 cells can improve the transfection efficiency.
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