Retrovirus-mediated gene targeting human Slingshot1L stable transfection of human bone marrow mesenchymal stem cell line.doc

 PAGE \* MERGEFORMAT 12 Retrovirus-mediated gene targeting human Slingshot1L stable transfection of human bone marrow mesenchymal stem cell line Of: Zhang Rui Wang Yan Ma Yingzhi Zhaodong Xu Yang Xufang Mu Li Yulin Abstract Objective To establish the gene over-expression of human bone marrow mesenchymal stem cells (hMSCs) stably transfected cell lines. Method combined with density gradient centrifugation separation of adherent culture, purified hMSCs. Application of liposome Slingshot 1L retroviral vector pLNCX Slingshot 1L transfected PA317 packaging cell, G418 selection, titer. Collection of viral supernatant infection of the 2nd generation hMSCs, G418 selection, the selection of positive clones were cultured. The results combined by density gradient centrifugation successfully separated adherent culture, purified hMSCs, application pLNCX Slingshot 1L recombinant retroviral vector-transfected PA317 packaging cells, obtained a titer of 5 108 CFU / L of viral supernatant. Collection of viral supernatant infection of hMSCs, obtained gene overexpression Slingshot 1L hMSCs. Conclusion Slingshot 1L recombinant retroviral vector, had successfully established the gene expression of hMSCs stably transfected cell lines. Keywords: retrovirus; stable transfection; stem cells Abstract 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 Slings