Cloning of bovine embryos from vitrified donor blastomeres

The use of cryopreserved in vitro produced bovine embryos as nuclear transfer donors was assessed. Day 4 or 5 morulae were vitrified and warmed using the open pulled straw method and used as donors for nuclear transfer. Although the proportion of morulae and blastocysts that developed from nuclear transfer embryos derived from day 5 vitrified embryos did not differ from that derived from fresh embryos (16.7 and 24.3%, respectively), development to blastocysts was reduced when vitrified donor cells were used (8.3 and 19.1%, respectively). Likewise, development to morulae and blastocysts was not different between nuclear transfer embryos derived from day 4 vitrified embryos allowed to recover for 24 h, and day 5 vitrified embryos allowed to recover for 1-2 h (27.7 and 15.6%, respectively), but the development to blastocysts was reduced when day 5 vitrified donor cells were used (23.2 and 10.0%, respectively). However, in nuclear transfer embryos derived from either day 4 vitrified or day 5 fresh donors, no differences were observed in development rates to morulae and blastocysts (34.3 and 36.3%, respectively) or to blastocysts alone (20.2 and 18.1%, respectively). Nor were there differences in the development rates of fresh or day 4 or day 5 vitrified in vitro produced (non-nuclear transfer) embryos (47.9, 51.0 and 35.5% developing to blastocysts at day 7, respectively). In vitro produced embryos and nuclear transfer embryos derived from day 4 vitrified or day 5 fresh donors were transferred to recipients at morula or blastocyst stage at day 6 or 7. The pregnancy rates were similar in both groups of nuclear transfer embryos, but higher in the control group consisting of in vitro produced embryos (47, 42 and 67%, respectively). In conclusion, if vitrified donor embryos are allowed to recover for 24 h after warming, their use in nuclear transfer results in similar efficiencies to those achieved with fresh embryos.