Mouse Induced Pluripotent Stem Cells Generated by piggyBac

Mouse somatic cells have been successfully reprogrammed into embryonic stem cell-like cells by enforced expression of defined transcription factors, and the reprogrammed cells are termed as induced pluripotent stem cells. Such cell reprogramming strategy is involved in gene transfection with retroviral, lentiviral, adenoviral and plasmid vectors. In addition, proteins and small molecules, synthetic modified mRNA and microRNA also have been used to efficiently generate mouse and human iPS. As a high efficient transposon, piggyBac has been used to generate induced pluripotent stem cells. In this study, piggyBac carrying four transcription factors called Oct4, Sox2, Klf4 and c-Myc has been employed to generate mouse induced pluripotent stem cells from murine embryonic fibroblasts, and the differentiation potential of the cells was assessed in vivo and in vitro. The reprogrammed cells showed positive staining for alkaline phosphatase and positive for OCT4, SSEA-1 and NANOG by immunofluorescent analyses. RT-PCR results showed that Oct4, Sox2, Klf4 and c-Myc was expressed in the reprogrammed cells. Immunofluorescent staining of AFP, BRACHYURY and NCAM1 indicated that the cells have the potential to differentiate spontaneously into all three germ layers in vitro. Teratomas formed after the cells subcutaneously injected into NOD-SCID mice. Chimeric blastocysts were developed when the cells were aggregated with mouse 8-cell embryos. These results suggested that mouse induced pluripotent stem cells can be generated from fibroblast cells by piggyBac transfection system. Such induced pluripotent stem cells are similar to mouse ES cells and have potential to differentiate into three germ layers in vitro and in vivo.