Comparison of the effects of using standard and simple portable CO 2 incubators on the in vitro developmental competence of bovine embryos reconstituted by somatic cell nuclear transfer

Abstract

In an attempt to determine the cultural factors that would improve cloning efficiency, we compared the effects of two incubation systems—a simple portable system and a standard CO 2 incubator—on the production of bovine embryos by electrofusion of quiescent fetal fibroblast nuclei to enucleated oocytes matured in vitro. While the temperature (38.5 °C) and CO 2 concentration (5%) were similar in both systems, the portable incubator operated in a vacuum of 300 mmHg and at an O 2 level of 8–10%, which is lower than the standard. Although there were no significant differences between the two systems in terms of in vitro oocyte maturation (MII stage), fusion rates, and the number of cells in Day 7 blastocysts, significantly higher proportions of nuclear-transferred oocytes cleaved ( P<0.05) and developed to the blastocyst stage ( P<0.01) in the portable incubator (70.5±0.6 and 36.1±1.4%, respectively) than in the standard incubator (64.1±3.2 and 23.5±1.4%, respectively). Following the transfer of six blastocysts from the portable incubator group to three recipients, survival rates on Days 60, 90, and 120 were 100, 66.7 and 33.3%, respectively. This relatively high early embryonic loss may be associated with multiple pregnancy complications or other abnormalities of placentation frequently observed in cloned embryos. Further studies using this portable incubator system are needed to determine the optimum levels of O 2, CO 2, and air pressure.