Isolation of Primordial Germ Cells from Differentiating Human Embryonic Stem Cells

Authors

  • Katarzyna Tilgner,

    1. North East Institute for Stem Cell Research, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
    2. Institute of Human Genetics, International Centre for Life, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
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  • Stuart P. Atkinson,

    1. North East Institute for Stem Cell Research, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
    2. Institute of Human Genetics, International Centre for Life, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
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  • Anna Golebiewska,

    1. North East Institute for Stem Cell Research, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
    2. Institute of Human Genetics, International Centre for Life, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
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  • Miodrag Stojković,

    1. Centro de Investigación Príncipe Felipe, Valencia, Spain
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  • Majlinda Lako,

    1. North East Institute for Stem Cell Research, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
    2. Institute of Human Genetics, International Centre for Life, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
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  • Lyle Armstrong

    Corresponding author
    1. North East Institute for Stem Cell Research, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
    2. Institute of Human Genetics, International Centre for Life, University of Newcastle upon Tyne, Newcastle upon Tyne, United Kingdom
    • Correspondence: Lyle Armstrong, BSc, Ph.D., International Centre for Life, Central Parkway, Newcastle upon Tyne NE1 3BZ, U.K. Telephone: 00-44-191-241-8695; Fax: 00-44-191-241-8666

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Abstract

Of all the cell types that can be obtained from the differentiation of embryonic stem cells, primordial germ cells are arguably the most fascinating, as they represent the in vitro completion of the reproductive cycle of the organism from which the embryonic stem cell line was derived. It is also possible to obtain these cells from embryos at an appropriate stage of development, but this process yields only small numbers that are not applicable to investigations of their epigenetic architecture. A considerable body of data has been generated from the differentiation of mouse embryonic stem cells to this cell type, but despite the demonstration of their presence in human embryoid bodies, there has been little progress toward methods of producing human primordial germ cells in useful numbers. We present here a robust protocol to differentiate two human embryonic stem cell lines (H9 and hES-NCL1) that maximizes the numbers of primordial germ cells that may be obtained using a simple fluorescence-activated cell sorting strategy for their isolation. These primordial germ cells demonstrate high-level expression of the germ cell-specific VASA gene and show removal of parental imprints and chromatin modification changes that support their primordial germ cell identity.

Disclosure of potential conflicts of interest is found at the end of this article.

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