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目的 了解含脑源性神经营养因子 (BDNF)基因的表达质粒PSVCEPBDNF CAT在培养肌母细胞中的表达情况及LipofectAMINE阳离子脂质体介导的转染效率。 方法 按每 2 μg表达质粒PSVCEPBDNF CAT由 2 0 μlLipofectAMINE包裹后转染至细胞浓度为 2× 1 0 8个 /L的3 .5cm培养皿培养的肌母细胞中 ,连续观察细胞形态 ,48h后用免疫细胞化学和免疫电镜的方法来检测其表达情况。结果 培养肌母细胞在转染 6h吞噬脂质体最显著 ,转染 48h后 ,40 %的肌母细胞胞质内有BDNF基因表达 ,免疫电镜结果证实表达在细胞质近胞膜处。结论 含BDNF微基因的表达质粒PSVCEPBDNF CAT可以在培养肌母细胞中表达 ,脂质体转染的方法简便、有效。
Objective To investigate the expression of brain-derived neurotrophic factor (BDNF) gene expression plasmid PSVCEPBDNF CAT in cultured myoblasts and LipofectAMINE cationic liposome-mediated transfection efficiency. Methods Each 2 μg expression plasmid PSVCEPBDNF CAT was encapsulated in 20 μl LipofectAMINE and then transfected into 3.5 × 10 8 cells / L cultured myoblasts with a cell concentration of 2 × 10 8 cells / L. The cell morphology was observed continuously. After 48 h, Immunocytochemistry and immunoelectron microscopy to detect the expression. Results The myoblasts were cultured in the presence of liposomes for 6 hours. After transfection for 48 hours, the expression of BDNF gene was found in the cytoplasm of 40% of the myoblasts. The results of immunoelectron microscopy showed that they were expressed in the cytoplasm near the cell membrane. Conclusion The expression plasmid PSVCEPBDNF CAT containing BDNF microgene can be expressed in cultured myoblasts. The liposome transfection method is simple and effective.