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BOR - Papers in Press, published online ahead of print March 3, 2004.
Biol Reprod 2004, 10.1095/biolreprod.104.027193
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Submitted January 2, 2004
Returned for revision February 4, 2004
Accepted March 3, 2004

Reproductive Technology


Cell Donor Influences Success of Producing Cattle by Somatic Cell Nuclear Transfer

A. M. Powell , N. C. Talbot , K. D. Wells , D. E. Kerr , V. G. Pursel , and R. J. Wall *

* To whom correspondence should be addressed. E-mail: bobwall{at}anri.barc.usda.gov.

Abstract
To assess sources of variation in nuclear transfer efficiency, bovine fetal fibroblasts (BFF), harvested from 6 Jersey fetuses, were cultured under various conditions. After transfection, frozen-thawed lung or muscle BFF donor cells were initially cultured in DMEM in 5% CO2 and air and some were transferred to MEM, with 5 or 20% O2 or 0.5 or 10% serum and G418 for 2 to 3 weeks. Selected clonal transfected fibroblasts were fused to enucleated oocytes. Fused couplets (n = 4007), activated with ionomycin and 6-dimethylaminopurine, yielded 927 blastocysts and 650 were transferred to 330 recipients. Fusion rate was influence by oxygen tension in a fetus dependent manner (P < 0.001). Blastocyst development was influenced in a number of ways. Hip fibroblast generated more blastocysts when cultured in MEM (P < 0.001). The influence of serum concentration was fetus dependent (P < 0.001) and exposing fibroblast to low oxygen was detrimental to blastocyst development (P < 0.001). Cells from 2 of the 6 fetuses produced embryos that maintained pregnancies to term, resulting in 8 viable calves. Pregnancy rates 56 d after transfer, for the 2 productive donor fetuses was at least double that of other recipients and may provide a fitness indicator of BFF cell sources for nuclear transfer. We conclude that a significant component in determining somatic cell nuclear transfer success is the source of the nuclear donor cells.

Key words: Embryo • Early development


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