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Biol Open. 2017 Apr 15;6(4):415-424. doi: 10.1242/bio.023473.

Reprogramming towards totipotency is greatly facilitated by synergistic effects of small molecules.

Author information

1
Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK masayasu@kais.kyoto-u.ac.jp kmiyamo@waka.kindai.ac.jp.
2
Laboratory of Molecular Developmental Biology, Graduate School of Biology-Oriented Science and Technology, Kindai University, Wakayama 649-6493, Japan.
3
Laboratory of Reproductive Biology, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan.
4
Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.
5
Institute of Advanced Technology, Kindai University, Wakayama 642-0017, Japan.
6
Laboratory of Reproductive Biology, Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan masayasu@kais.kyoto-u.ac.jp kmiyamo@waka.kindai.ac.jp.

Abstract

Animal cloning has been achieved in many species by transplanting differentiated cell nuclei to unfertilized oocytes. However, the low efficiencies of cloning have remained an unresolved issue. Here we find that the combination of two small molecules, trichostatin A (TSA) and vitamin C (VC), under culture condition with bovine serum albumin deionized by ion-exchange resins, dramatically improves the cloning efficiency in mice and 15% of cloned embryos develop to term by means of somatic cell nuclear transfer (SCNT). The improvement was not observed by adding the non-treated, rather than deionized, bovine serum. RNA-seq analyses of SCNT embryos at the two-cell stage revealed that the treatment with TSA and VC resulted in the upregulated expression of previously identified reprogramming-resistant genes. Moreover, the expression of early-embryo-specific retroelements was upregulated by the TSA and VC treatment. The enhanced gene expression was relevant to the VC-mediated reduction of histone H3 lysine 9 methylation in SCNT embryos. Our study thus shows a simply applicable method to greatly improve mouse cloning efficiency, and furthers our understanding of how somatic nuclei acquire totipotency.

KEYWORDS:

Epigenetic modification; Mouse; Nuclear transfer; Reprogramming

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