Nuclear Transplantation in Early Pig Embryosl

Nuclear transfer was evaluated in early porcine embryos. Pronuclear stage embryos were centr(fuged, treated with cytoskeletal inhibitors, and subsequently enuclewed. Pronuclei containing karyoplasts were placed in the perivitelline space of the enucleazed zygote and fused to the enucleated zygote with electrofusion. The resulting pronuclear exchange embryos were either monitored for cleavage in vitro (9/13 cleaved and contained 2 nuclei after 24 h, 69%) or for in vivo development. In vivo development after 3 days resulted in 14/15 (93%) of the embryos transferred cleaving to the (cid:18)4-cell stage and after 7 days 6/16 (38%) reaching the expanded blastocys: stage. A total of 56 pronuclear exchange embryos were allowed to go to term, and 7 piglets were born. A similar manipulation procedure was used to transfer 2-, 4- or 8-cell nuclei to enucleated, activated meiotic metaphase II oocytes. Enucleation was effective in 74% (36/49) of the contemporary oocy:es. Activation was successful in 81% (37/46) of nonmanipulated but pulsed oocytes versus 13% (4/31) of control oocytes (p<O.Ol). After 6 days in vivo, 9% (1/11) of the 2-cell nuclei, 8% (7/83) of the 4-cell nuclei, and 19% (11/57) of the 8-cell nuclei transferred to enucleated, activated meiotic metaphase 11 oocytes resulted in development to the compact morula or blastocys: stage (pczO.OI). A total of 88 nuclear transfer embryos were transferred to recipient gills for continued development. A single piglet was born after the transfer of a 4-cell nucleus zoan enucleated, activated metaphase 11 oocy:e and subsequent in vivo development. Therefore 4-cell nuclei are capable of directing development to term after transfer to an enucleated, activated meiotic metaphase I! oocyte.

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