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BOR - Papers in Press, published online ahead of print October 4, 2006.
Biol Reprod 2006, 10.1095/biolreprod.106.051383
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Submitted February 2, 2006
Returned for revision March 2, 2006
Accepted September 19, 2006

Embryo


Epigenetic Marks in Cloned Rhesus Monkey Embryos: Comparison with Counterparts Produced In Vitro

Jifeng Yang , Shihua Yang , Nathalie Beaujean , Yuyu Niu , Xiechao He , Yunhua Xie , Xianghui Tang , Liu Wang , Qi Zhou , and Weizhi Ji *

* To whom correspondence should be addressed. E-mail: wji{at}mail.kiz.ac.cn.

Abstract
Up to now, no primate animals have been successfully cloned to birth with somatic cells nuclear transfer (SCNT) and little is known about the molecular events occurring in the reconstructed embryos during preimplantation development. In many SCNT cases, epigenetic reprogramming of the donor nuclei after transfer into enucleated oocytes was hypothesized to be crucial in the reestablishment of embryonic totipotency. In this study, we focused on two major epigenetic marks, DNA methylation and histone H3 lysine 9 (H3K9) acetylation, examined by indirect immunofluorescence and confocal laser scanning microscopy. During preimplantation development, 67% of 2-cell and 50% of 8-cell cloned embryos showed higher DNA methylation levels than their IVF counterparts which undergo gradual demethylation until the early morula stage. Moreover, whereas an asymmetric distribution of DNA methylation was established in IVF blastocyst with a lower methylation level in the inner cell mass (ICM) than in the trophectoderm, in most cloned blastocysts ICM cells maintained a high degree of methylation. Finally, two donor cells lines (S11 and S1-04) that showed a higher level of H3K9 acetylation supported more blastocyst formation after nuclear transfer than the other cell line (S1-03) with relative low level of acetylation staining. In conclusion, we propose that abnormal DNA methylation patterns contributes to the poor quality of cloned preimplantation embryos and may be one of the obstacles to successful cloning in primates.

Key words: Embryo • Early development • In vitro fertilization


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