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Hum. Reprod. Advance Access published online on June 22, 2006
Human Reproduction, doi:10.1093/humrep/del216
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© The Author 2006. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions@oxfordjournals.org
Received April 1, 2006
Revised May 3, 2006
Accepted May 12, 2006

Article

A comparative approach to somatic cell nuclear transfer in the rhesus monkey

Q. Zhou 1 *, S.H. Yang 2 *, C.H. Ding 2 *, X.C. He 3, Y.H. Xie 3, T.B. Hildebrandt 4, S.M. Mitalipov 5, X.H. Tang 3, D.P. Wolf 5 *, and W.Z. Ji 3 *

1 State Key Lab of Reproductive Biology, Institute of Zoology, The Chinese Academy of Sciences, Beijing; Kunming Primate Research Center and Kunming Institute of Zoology, The Chinese Academy of Sciences, Yunnan, China
2 Kunming Primate Research Center and Kunming Institute of Zoology, The Chinese Academy of Sciences, Yunnan, China; The same contribution as first author
3 Kunming Primate Research Center and Kunming Institute of Zoology, The Chinese Academy of Sciences, Yunnan, China
4 Institute for Zoo and Wildlife Research (IZW), Berlin, Germany
5 Oregon National Primate Research Center, Beaverton, OR, USA

* To whom correspondence should be addressed.
Q. Zhou, E-mail: qzhou{at}ioz.ac.cn
D.P. Wolf, E-mail: wolfd{at}ohsu.edu
W.Z. Ji, E-mail: wji{at}mail.kiz.ac.cn


  Abstract

BACKGROUND: Despite the potential utility of primate somatic cell nuclear transfer (SCNT) to biomedical research and to the production of autologous embryonic stem (ES) cells for cell- or tissue-based therapy, a reliable method for SCNT is not yet available. Employing the rhesus monkey as a clinically relevant animal model, we have compared a conventional electrofusion method for SCNT with a one-step micromanipulation (OSM) method. METHODS: A prospective, randomized trial was conducted using only oocytes that were mature [metaphase II (MII)] at collection and a fibroblast-like cell line as nuclear donor cells (fetal fibroblasts). The embryos produced were characterized for in vitro developmental potential, cell number, karyotype and expression of nuclear mitotic apparatus (NuMA) and OCT-4. RESULTS: An in vitro blastocyst development rate of 24.4% was achieved with the OSM method, significantly higher than the 12.2% obtained following electrofusion. SCNT-produced embryos expressed normal karyotypes, cell numbers and NuMA and OCT-4 proteins in most cases. SCNT with male nuclear donor cells resulted in the production of male, SCNT blastocysts, eliminating the possibility of a parthenogenetic origin. Of the four fibroblast cell lines tested as nuclear donor cells, two supported the routine production of blastocysts following SCNT. CONCLUSIONS: The application of a modified SCNT technique (OSM) followed by embryo culture in hamster embryo culture medium-10 (HECM-10) allows, for the first time, the routine production of SCNT blastocysts, most of which appear normal by immunochemical, cytochemical and in vitro developmental criteria. These embryos will provide a resource for isolating ES cells and for studies of nuclear reprogramming by monkey cytoplasts.

Keywords: rhesus monkey/somatic cell nuclei transfer/therapeutic cloning.

* The same contribution as first author.


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