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Development. 2014 Oct;141(20):3842-7. doi: 10.1242/dev.110726. Epub 2014 Sep 24.

Induction of the G2/M transition stabilizes haploid embryonic stem cells.

Author information

1
Department of Epigenetics, Medical Research Institute, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
2
Department of Epigenetics, Medical Research Institute, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan Global Center of Excellence Program for International Research Center for Molecular Science in Tooth and Bone Diseases, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
3
Center for Experimental Animals, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan.
4
School of Health Sciences, Tokai University, Bohseidai, Isehara, Kanagawa 259-1193, Japan tkanekoi@is.icc.u-tokai.ac.jp fishino.epgn@mri.tmd.ac.jp.
5
Department of Epigenetics, Medical Research Institute, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan Global Center of Excellence Program for International Research Center for Molecular Science in Tooth and Bone Diseases, Tokyo Medical and Dental University (TMDU), 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8510, Japan tkanekoi@is.icc.u-tokai.ac.jp fishino.epgn@mri.tmd.ac.jp.

Abstract

The recent successful establishment of mouse parthenogenetic haploid embryonic stem cells (phESCs) and androgenetic haploid ESCs (ahESCs) has stimulated genetic research not only in vitro but also in vivo because of the germline competence of these cell lines. However, it is difficult to maintain the haploid status over time without a frequent sorting of the G1 phase haploid ESCs by fluorescence-activated cell sorting (FACS) at short intervals, because haploid cells tend to readily self-diploidize. To overcome this spontaneous diploid conversion, we developed a phESC culture condition using a small molecular inhibitor of Wee1 kinase to regulate the cell cycle by accelerating the G2/M phase transition and preventing re-entry into extra G1/S phase. Here, we demonstrate that, under this condition, phESCs maintained the haploid status for at least 4 weeks without the need for FACS. This method will greatly enhance the availability of these cells for genetic screening.

KEYWORDS:

Cell cycle; Haploid embryonic stem cells; Mouse; Parthenogenesis

PMID:
25252944
DOI:
10.1242/dev.110726
[Indexed for MEDLINE]
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