CAJ | 학술논문

目的建立过表达该基因的人骨髓间充质干细胞(hMSCs)稳定转染细胞系.方法采用密度梯度离心法联合贴壁培养法分离、纯化hMSCs .应用脂质体法将Slingshot-1L 重组逆转录病毒载体pLNCX-Slingshot-1L转染PA317包装细胞,G418筛选,滴度测定.收集病毒上清感染第2代hMSCs,G418筛选,挑选阳性克隆,扩增培养.结果采用密度梯度离心法联合贴壁培养法成功分离、纯化了hMSCs,应用pLNCX-Slingshot-1L重组逆转录病毒载体转染PA317包装细胞,获得滴度为5×10~8 CFU /L的病毒上清.收集病毒上清感染hMSCs,获得了过表达Slingshot-1L基因的hMSCs.结论应用Slingshot-1L重组逆转录病毒载体,成功建立了过表达该基因的hMSCs稳定转染细胞系.
목적 건립과표체해기인적인골수간충질간세포(hMSCs)은정전염세포계.방법 채용밀도제도리심법연합첩벽배양법분리、순화hMSCs .응용지질체법장Slingshot-1L 중조역전록병독재체pLNCX-Slingshot-1L전염PA317포장세포,G418사선,적도측정.수집병독상청감염제2대hMSCs,G418사선,도선양성극륭,확증배양.결과 채용밀도제도리심법연합첩벽배양법성공분리、순화료hMSCs,응용pLNCX-Slingshot-1L중조역전록병독재체전염PA317포장세포,획득적도위5×10~8 CFU /L적병독상청.수집병독상청감염hMSCs,획득료과표체Slingshot-1L기인적hMSCs.결론 응용Slingshot-1L중조역전록병독재체,성공건립료과표체해기인적hMSCs은정전염세포계.
Objective To establish stable transfected human marrow mesenchymal stem cells (hMSCs) line by retroviral vector encoding Slingshot-1Lto abtain viral supernatant with high viral titers. Methods hMSCs were isolated and purified from the bone marrow of human by density gradient centrifugation and adhering to the culture dishes. The plasmid pLNCX-Slingshot-1L was transfected into packaging cell PA317 by lipofectamine. The transfectants were selected by G418 and viral titers were analyzed. Virals supernatant were collected to infect hMSCs in the second passage, selected by G418. Results Virals supernatant with high viral titers were obtained by retroviral vector encoding Slingshot-1L, virals supernatant were collected to infect hMSCs, and stable transfected hMSCs line was established. Conclusions The stable transfected hMSCs line over express Slingshot-1L mediated by retroviral was established successfully.