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In vitro generation of germ cells from murine embryonic stem cells

Nature Protocols volume 1, pages 2026–2036 (2006)Cite this article

Abstract

The demonstration of germ cell and haploid gamete development from embryonic stem cells (ESCs) in vitro has engendered a unique set of possibilities for the study of germ cell development and the associated epigenetic phenomenon. The process of embryoid body (EB) differentiation, like teratoma formation, signifies a spontaneous differentiation of ESCs into cells of all three germ layers, and it is from these differentiating aggregates of cells that putative primordial germ cells (PGCs) and more mature gametes can be identified and isolated. The differentiation system presented here requires the differentiation of murine ESCs into EBs and the subsequent isolation of PGCs as well as haploid male gametes from EBs at various stages of differentiation. It serves as a platform for studying the poorly understood process of germ cell allocation, imprint erasure and gamete formation, with 4–6 weeks being required to isolate PGCs as well as haploid cells.

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Figure 1: The stages of germ cell and haploid gamete selection from ESC differentiation.
Figure 2: EGC clones demonstrate imprint erasure in a time-dependent manner during EB differentiation.
Figure 3: RT–PCR of germ cell-specific genes during EB differentiation.
Figure 4: Germ cell formation occurs in a time-dependent manner during EB differentiation.
Figure 5: EBs generate haploid male gametes that can be enriched by FE-J1 staining.

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Authors and Affiliations

  1. Graduate Program of Biological and Biomedical Sciences, Harvard Medical School, Boston, Massachusetts, USA

    Jason A West

  2. Division of Pediatric Hematology/Oncology, Children's Hospital Boston and Dana Farber Cancer Institute, Boston, 02115, Massachusetts, USA

    Jason A West, In-Hyun Park & George Q Daley

  3. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Jason A West, In-Hyun Park & George Q Daley

  4. Harvard Stem Cell Institute, Boston, 02115, Massachusetts, USA

    Jason A West, In-Hyun Park & George Q Daley

  5. Center for Regenerative Medicine, Massachusetts General Hospital, Harvard Stem Cell Institute, Boston, 02114, Massachusetts, USA

    Niels Geijsen

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Correspondence to George Q Daley or Niels Geijsen.

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West, J., Park, IH., Daley, G. et al. In vitro generation of germ cells from murine embryonic stem cells. Nat Protoc 1, 2026–2036 (2006). https://doi.org/10.1038/nprot.2006.303

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