Jiang, Yan, Kelly, Richard D. W., Peters, Amy, Fulka, Helena, Dickinson, Adam, Mitchell, Daniel A. and St. John, Justin C. (2011) Interspecies somatic cell nuclear transfer is dependent on compatible mitochondrial DNA and reprogramming factors. PL o S One, Vol.6 (No.4). e14805. doi:10.1371/journal.pone.0014805 ISSN 1932-6203.

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Abstract

Interspecies somatic cell nuclear transfer (iSCNT) involves the transfer of a nucleus or cell from one species into the
cytoplasm of an enucleated oocyte from another. Once activated, reconstructed oocytes can be cultured in vitro to
blastocyst, the final stage of preimplantation development. However, they often arrest during the early stages of
preimplantation development; fail to reprogramme the somatic nucleus; and eliminate the accompanying donor cell’s
mitochondrial DNA (mtDNA) in favour of the recipient oocyte’s genetically more divergent population. This last point has
consequences for the production of ATP by the electron transfer chain, which is encoded by nuclear and mtDNA. Using a
murine-porcine interspecies model, we investigated the importance of nuclear-cytoplasmic compatibility on successful
development. Initially, we transferred murine fetal fibroblasts into enucleated porcine oocytes, which resulted in extremely
low blastocyst rates (0.48%); and failure to replicate nuclear DNA and express Oct-4, the key marker of reprogramming.
Using allele specific-PCR, we detected peak levels of murine mtDNA at 0.1460.055% of total mtDNA at the 2-cell embryo
stage and then at ever-decreasing levels to the blastocyst stage (,0.001%). Furthermore, these embryos had an overall
mtDNA profile similar to porcine embryos. We then depleted porcine oocytes of their mtDNA using 10 mM 29,39-
dideoxycytidine and transferred murine somatic cells along with murine embryonic stem cell extract, which expressed key
pluripotent genes associated with reprogramming and contained mitochondria, into these oocytes. Blastocyst rates
increased significantly (3.38%) compared to embryos generated from non-supplemented oocytes (P,0.01). They also had
significantly more murine mtDNA at the 2-cell stage than the non-supplemented embryos, which was maintained
throughout early preimplantation development. At later stages, these embryos possessed 49.9962.97% murine mtDNA.
They also exhibited an mtDNA profile similar to murine preimplantation embryos. Overall, these data demonstrate that the
addition of species compatible mtDNA and reprogramming factors improves developmental outcomes for iSCNT embryos.

Item Type:	Journal Article
Subjects:	Q Science > QH Natural history
Divisions:	Faculty of Science, Engineering and Medicine > Medicine > Warwick Medical School
Library of Congress Subject Headings (LCSH):	Cell nuclei -- Transplantation, Somatic cells, Embryonic stem cells, Mitochondrial DNA
Journal or Publication Title:	PL o S One
Publisher:	Public Library of Science
ISSN:	1932-6203
Official Date:	27 April 2011
Dates:	Date Event 27 April 2011 Published
Volume:	Vol.6
Number:	No.4
Page Range:	e14805
DOI:	10.1371/journal.pone.0014805
Status:	Peer Reviewed
Access rights to Published version:	Restricted or Subscription Access
Funder:	Medical Research Council (Great Britain) (MRC)
Grant number:	G0600273 (MRC)

Data sourced from Thomson Reuters' Web of Knowledge

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