Production of Cloned Bighorn Sheep Embryos Using ISCNT via HMC from Domestic Sheep Oocytes Treated with Resveratrol During IVM

The aim of this study was to treat sheep oocytes with resveratrol during in vitro maturation (IVM) and use them as cytoplasts in ISCNT via handmade cloning (HMC), evaluating the effect of resveratrol on the in vitro development of cloned Mexican bighorn sheep embryos. Post mortem skin fibroblasts from an adult male specimen were frozen for 8 years, thawed, and reseeded for eight cell passages. For IVM, Ovis aries oocytes were treated with 0, 0.5, or 1.0 µM resveratrol. Matured oocytes were manually enucleated, and triplets (O. aries cytoplast–Ovis canadensis mexicana karyoplast–O. aries cytoplast) were formed and electrically fused. The reconstructed embryos were chemically activated and cultured for in vitro development (IVD). The IVM rate was 81.8 ± 10.4% for CG, 81.9 ± 6.7% for EG1, and 76.3 ± 7.7% for EG2, with no significant differences between groups. For IVD, EG1 showed a statistically significant increase (p < 0.05) in the blastocyst rate (31 ± 12.0%) and a statistically significant decrease in the fragmented embryo rate (25 ± 10.4) when compared with the other groups. It was concluded that better rates of cloned bighorn sheep blastocysts could be obtained in ISCNT via HMC when fusing O. aries oocytes supplemented with resveratrol during IVM with post mortem adult male O. c. mexicana fibroblasts that had been cryopreserved for 8 years.