Embryonic Stem Cells/Induced Pluripotent Stem Cells

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Expression of GFP Under the Control of the RNA Helicase VASA Permits Fluorescence-Activated Cell Sorting Isolation of Human Primordial Germ Cells†‡§

  1. Katarzyna Tilgner1,2,
  2. Stuart P. Atkinson1,2,
  3. Sun Yung1,2,
  4. Anna Golebiewska1,2,
  5. Miodrag Stojkovic3,
  6. Ruben Moreno3,
  7. Majlinda Lako1,2,
  8. Lyle Armstrong1,2,¶,*

Article first published online: 23 NOV 2009

DOI: 10.1002/stem.263

Issue

STEM CELLS

STEM CELLS

Volume 28, Issue 1, pages 84–92, January 2010

Additional Information

How to Cite

Tilgner, K., Atkinson, S. P., Yung, S., Golebiewska, A., Stojkovic, M., Moreno, R., Lako, M. and Armstrong, L. (2010), Expression of GFP Under the Control of the RNA Helicase VASA Permits Fluorescence-Activated Cell Sorting Isolation of Human Primordial Germ Cells. STEM CELLS, 28: 84–92. doi: 10.1002/stem.263

Author Information

  1. 1

    North East Stem Cell Institute and International Centre for Life, NE1 3BZ, United Kingdom

  2. 2

    Institute of Human Genetics, Newcastle University, International Centre for Life, NE1 3BZ, United Kingdom

  3. 3

    Centro de Investigación Príncipe Felipe C/E.P. Avda. Autopista del Saler, 16-3 (junto Oceanográfico), 46012 Valencia, Spain

  1. Telephone: 00 44 191 241 8695; Fax: 00 44 191 241 8666

*International Centre for Life, Central Parkway, Newcastle upon Tyne NE1 3BZ, U.K

  1. †

    Author contributions: K.T., S.P.A., S.Y., and A.G.: Collection and assembly of data; M.S.: Manuscript writing; R.M.: Manuscript writing and fund raising; M.L. and L.A.: Conception and design, manuscript writing, and fund raising.

  2. ‡

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

  3. §

    First published online in STEM CELLS EXPRESS November 23, 2009.

Publication History

  1. Issue published online: 12 JAN 2010
  2. Article first published online: 23 NOV 2009
  3. Accepted manuscript online: 23 NOV 2009 12:00AM EST
  4. Manuscript Accepted: 11 NOV 2009
  5. Manuscript Received: 20 MAR 2009

Funded by

  • Conselleria de Sanidad (Generalitat Valenciana)
  • Instituto de Salud Carlos III (Ministry of Science and Innovation)

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Keywords:

  • Embryonic Stem Cells;
  • Primordial Germ Cells;
  • Meiosis;
  • VASA

Abstract

The isolation of significant numbers of human primordial germ cells at several developmental stages is important for investigations of the mechanisms by which they are able to undergo epigenetic reprogramming. Only small numbers of these cells can be obtained from embryos of appropriate developmental stages, so the differentiation of human embryonic stem cells is essential to obtain sufficient numbers of primordial germ cells to permit epigenetic examination. Despite progress in the enrichment of human primordial germ cells using fluorescence-activated cell sorting (FACS), there is still no definitive marker of the germ cell phenotype. Expression of the widely conserved RNA helicase VASA is restricted to germline cells, but in contrast to species such as Mus musculus in which reporter constructs expressing green fluorescent protein (GFP) under the control of a Vasa promoter have been developed, such reporter systems are lacking in human in vitro models. We report here the generation and characterization of human embryonic stem cell lines stably carrying a VASA-pEGFP-1 reporter construct that expresses GFP in a population of differentiating human embryonic stem cells that show expression of characteristic markers of primordial germ cells. This population shows a different pattern of chromatin modifications to those obtained by FACS enrichment of Stage Specific Antigen one expressing cells in our previous publication. STEM CELLS 2010;28:84–92

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