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Objective: ectopic expression of certain transcription factors can reprogramming somatic cells into induced pluripotent stem cells ( iPSCs ) can avoid the ethical issues brought about by the ES cells and reduce immune rejection , to the autologous stem cell gene therapy the clinical application provides the possibility of genetic disease in vitro studies provide a good platform . Methods: There are a variety of methods can induce iPS cells, including the use of a variety of viral vectors, the purified recombinant protein, modified RNA molecules . Also found that a variety of small molecular compounds can also facilitate the generation of iPS , Vc can accelerate the change in level of gene expression , and promote precursor iPS cells, the iPS cells into complete reprogramming ; VPA in the reprogramming process , can be raised ES specific gene expression, down specific gene expression in MEF . In the present study , we used a retrovirus-mediated transfection system the transcription factor Oct4, Sox2, c-Myc , and Klf4 4 is introduced into a human skin fibroblast cells ( HDF ) , while adding Vc and VPA during induction and, ultimately, the iPS cells . Then, through TRA-1-60/TRA-1-81 live cell staining , mechanical pick positive cloning subcultured . Taken morphology , alkaline phosphatase staining , and a stem cell surface the pluripotency marker staining method for preliminary identification of the passaged clone . Results: (1 ) using liposome Lipofectamine2000 retroviral vectors and packaging plasmids co-transfected 293T cells, and a higher quality can be obtained retrovirus ; (2) Cloning of induced morphological similarity with hES cells , the outer gene GFP expression of after living cells staining found its TRA-1-60/TRA-1-81 expression , picked the clone passaged after AP staining results were positive ES cell surface landmark immunofluorescence tested positive . Spread P7 generation , still has the ES - like colonies form . Conclusion : The use of retroviral system to human skin fibroblast cells reprogrammed iPS cells express pluripotency surface markers was identified .
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