Production of mice using iPS cells and tetraploid complementation.

State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, China.
Nature Protocol (Impact Factor: 8.36). 04/2010; 5(5):963-71. DOI: 10.1038/nprot.2010.61
Source: PubMed

ABSTRACT Induced pluripotent stem cells (iPSCs) are considered to be an attractive alternative to embryonic stem cells (ESCs) and may provide great potential for clinical applications in regenerative medicine. Although possessing characteristics similar to ESCs, the true pluripotency of these newly studied iPSCs was not known because none of the previously developed iPSCs passed the tetraploid complementation assay, which is regarded as the most stringent test for pluripotency. We have recently shown that by modifying some of the culture conditions for inducing iPSCs, we were able to generate cell lines of high pluripotency, resulting in the production of live-born, fertile animals through tetraploid complementation. In this paper, we describe details of our methods of generating iPS cell lines and subsequently producing full-term live animals through the tetraploid complementation assay; the procedure can be completed within 2 months.

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    ABSTRACT: Objectives Recently, pluripotency of induced pluripotent stem (iPS) cells has been displayed after producing adult mice, in tetraploid complementation assays. These studies lead us to the last piece of the puzzle for reprogramming somatic cells into fully pluripotent cells which function as embryonic stem cells in most applications. However, in all of previous studies, skin fibroblasts were used as the starting population for reprogramming, raising questions as to whether the pluripotency of the iPS cells was dependent on the particular starting cell type.Materials and methodsOur iPS cell lines were prepared from murine adipose stem cells (ASCs). Their multi-potency was first tested by teratoma formation in nude mice. Then, tetraploid complementation was performed to generate progeny from them.ResultsWe succeeded to the birth of viable and fertile adult mice derived entirely from reprogrammed ASC, indicating cell types other than fibroblasts can also be restored to the embryonic level of pluripotency.Conclusions We also directed differentiation of iPS cells into chondrocytes, thus adipose-derived iPS cells can be used as models to study chondrogenic differentiation and cartilage regeneration.
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