Induction of pluripotent stem cells from fibroblast cultures

Abstract

Clinical application of embryonic stem (ES) cells faces difficulties regarding use of embryos, as well as tissue rejection after implantation. One way to circumvent these issues is to generate pluripotent stem cells directly from somatic cells. Somatic cells can be reprogrammed to an embryonic-like state by the injection of a nucleus into an enucleated oocyte or by fusion with ES cells. However, little is known about the mechanisms underlying these processes. We have recently shown that the combination of four transcription factors can generate ES-like pluripotent stem cells directly from mouse fibroblast cultures. The cells, named induced pluripotent stem (iPS) cells, can be differentiated into three germ layers and committed to chimeric mice. Here we describe detailed methods and tips for the generation of iPS cells.

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Figure 1: Schematic diagram of induced pluripotent stem (iPS) cell generation.
Figure 2: Morphologies of mouse fibroblasts used for induced pluripotent stem (iPS) cell induction.
Figure 3: Retroviral transfection system for induced pluripotent stem (iPS) cell generation.
Figure 4: Induced pluripotent stem (iPS) cells from mouse fibroblasts.

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Acknowledgements

We are grateful to Dr. Yoshimi Tokuzawa for earlier works that were fundamental for our experiments. We also thank Tomoko Ichisaka, Megumi Narita, Yumi Ohuchi, Rie Kato and Ryoko Iyama for technical and administrative supports, and Dr. Toshio Kitamura for pMXs retroviral vectors and Plat-E cells.

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Correspondence to Shinya Yamanaka.

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Takahashi, K., Okita, K., Nakagawa, M. et al. Induction of pluripotent stem cells from fibroblast cultures. Nat Protoc 2, 3081–3089 (2007). https://doi.org/10.1038/nprot.2007.418

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