OOCYTE ACTIVATION BY STRONTIUM IN THE PRESENCE OF CALCIUM SUPPORTS FULL TERM DEVELOPMENT OF SOMATIC CELL CLONED EMBRYOS

Abstract

Without using sperm, artificial oocyte activation is essential for the current assisted reproductive technologies, particularly somatic cell nuclear transfer and round spermatid injection. Strontium has been widely used as an activator of oocytes especially in mouse, by which efficient oocyte activation requires calcium-free medium but not culture media since they normally contain calcium. In this study, we considered whether strontium with chelated calcium in culture media is enough to efficiently activate oocytes. Ethylene glycol-bis (beta-aminoethyl ether) - N, N, N', N'-tetraacetic acid (EGTA) were added to three standard culture media (CZB, M16 and KSOM) for mouse embryos because they contain EGTA which preferentially binds calcium ion rather than strontium ion. After optimizing EGTA at 2–5 mM along with 5mM strontium in either CZB, M16 or KSOM, more than 80% of available oocytes were parthenogenetically activated, which was as efficient as standard conditions of strontium in a Ca-free medium. Further, it was demonstrated that this activation method can support full-term development of cloned embryos using trichostatin A with comparable success rates. Thus, addition of EGTA into culture media can easily make activation media without additional preparation of Ca-free medium. (poster)