In Vitro Development and Production of Offspring from Bovine Nuclear Transfer Embryos Cultured in a Serum-free Medium

Abstract

Newly developed serum-free media (IVD101 in the absence of bovine cumulus/granulosa cell (BCGC) coculture and IVMD101 in the presence of BCGC coculture) have recently been found to improve the yield and quality of bovine blastocysts from in vitro matured and fertilized oocytes [1]. The present study was undertaken with two objectives: first, to examine whether bovine nuclear transfer embryos could develop to the blastocyst stage when cultured in the serum-free media (IVD101 and IVMD101): second, to confirm the potency of the nuclear transfer blastocysts to develop to viable offspring after transfer to recipient cows. In vivo-derived embryos were used as donor nuclei and in vitro-derived oocytes were used as recipient oocytes for nuclear transfer. Recipient oocytes at the second metaphase were enucleated and the fusion of donor blastomere with the recipient oocytes was carried out by electrostimulation. The nuclear transfer embryos cultured in IVMD101 with BCGC coculture and IVD101 without BCGC coculture resulted in 20.4% and 27.5% blastocyst formation rate, respectively, which were similar to that (23.4%) of nuclear transfer embryos cultured in a conventional serum-supplemented medium (CR1aa+5% calf serum; CS) with BCGC coculture. Transfer of the nuclear transfer blastocysts developed in IVMD101, IVD101 and CR1aa+5%CS resulted in one pregnancy (1/7), two pregnancies (2/6), and two pregnancies (2/5), respectively, when examined on Day 40. The corresponding numbers of normal live calves born were 1, 1, and 1. Our results indicate that the two different serum-free media may be useful for the in vitro development of nuclear transfer embryos and the subsequent production of normal calves after transfer to recipient cows.