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Isolation and cultivation of stem cells from adult mouse testes

Abstract

The successful isolation and cultivation of spermatogonial stem cells (SSCs) as well as induction of SSCs into pluripotent stem cells will allow us to study their biological characteristics and their applications in therapeutic approaches. Here we provide step-by-step procedures on the basis of previous work in our laboratory for: the isolation of testicular cells from adolescent mice by a modified enzymatic procedure; the enrichment of undifferentiated spermatogonia by laminin selection or genetic selection using Stra8-EGFP (enhanced green fluorescent protein) transgenic mice; the cultivation and conversion of undifferentiated spermatogonia into embryonic stem-like cells, so-called multipotent adult germline stem cells (maGSCs); and characterization of these cells. Normally, it will take about 16 weeks to obtain stable maGSC lines starting from the isolation of testicular cells.

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Figure 1
Figure 2: Phase-contrast images of spermatogonial colonies and ES-like colony.
Figure 3: Regeneration of spermatogenesis by Stra8-EGFP+ SSCs.
Figure 4: Pluripotency analysis of maGSCs derived from mouse SSCs.
Figure 5: Differentiation of maGSCs in vitro and in vivo.

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Acknowledgements

We are grateful to Anke Cierpka and all members of the Guan and Hasenfuss lab for their contributions to this work. We thank Jan Streckfuß for the excellent record of the video. The methods described in this article were developed in the Guan and Hasenfuss lab in collaboration with W Engel and K Nayernia (Institute of Human Genetics, Georg-August-University of Göttingen, Germany). Our studies are supported by a Heidenreich von Siebold-Program 2006 grant from Georg-August-University of Göttingen to K.G., a BMBF grant to G.H. (01GN0601), a 'Forschungs-und Berufungspool (Kapitel 06 08 TG 74)' grant of Ministry for Science and Culture of Lower Saxony (G.H.), and a DFG grant (Klinische Forschergruppe KFO 155) to K.G., W.E. and G.H.

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Correspondence to Kaomei Guan or Gerd Hasenfuss.

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Guan, K., Wolf, F., Becker, A. et al. Isolation and cultivation of stem cells from adult mouse testes. Nat Protoc 4, 143–154 (2009). https://doi.org/10.1038/nprot.2008.242

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