Rapsyn重组质粒转染小鼠骨髓间充质干细胞

目的探讨采用脂质体介导的基因转移技术将重组pEGFP-N1/Rapsyn质粒转染至小鼠骨髓间充质干细胞(BMSCs)的可能性,并观察Rapsyn蛋白在靶细胞中的表达。方法采用全骨髓贴壁培养法分离纯化C57BL/6小鼠的BMSCs,在体外进行扩增、传代。取第3代细胞,采用脂质体介导的基因转移技术将重组pEGFP-N1/Rapsyn质粒转染至BMSCs中,并置荧光显微镜下观察转染结果;采用Western blot法检测靶细胞中Rapsyn蛋白的表达。结果 BMSCs经转染后24h可在荧光显微镜下观察到绿色荧光,转染pEGFP-N1/Rapsyn质粒的BMSCs可稳定表达Rapsyn蛋白。结论利用脂质体介导法可将Rapsyn基因转染至BMSCs中,并能稳定表达Rapsyn蛋白。

Study on bone marrow mesenchymal stem cells transfected by eukaryotic expression vector carrying rapsyn gene

Objective To explore the possibility of transfecting mouse bone marrow mesenchymal stem cells(BMSCs) by using pEGFP-N1/Rapsyn eukaryotic expressing plasmid,and to detect rapsyn protein expression in target cells.Methods C57BL/6 mouse BMSCs were isolated and purified by attachment culture method,in vitro amplification and passage were conducted.The pEGFP-N1/Rapsyn expressing plasmid was transfected into the 3rd passage BMSCs by liposome-mediated gene transfer technique,the green fluorescence was observed under fluorescence microscope,and rapsyn protein expression was detected by western blot.Results 24 h after transfection,green fluorescence can be observed under a fluorescence microscope,and rapsyn protein in the transfected group was detected by western blot.Conclusions The rapsyn gene can be successfully transfected into mouse BMSCs and keep on expressing.