Studies on the Porcine Oocytes Activation Regimed for Nuclear Transfer and Development Following Co-culture of Nuclear Transferred Embryos

Kim, Sang-keun; Rhee, Man-hwi

Korean Journal of Veterinary Research 2002;42(1):29-33.

Sang-keun Kim¹, Man-hwi Rhee²

¹College of Veterinary Medicine, Chungnam National University
²Dept. of Neurophysiology, Washington University

핵 이식을 위한 돼지난자의 활성화 및 핵 이식 배와 공배양에 따른 체외발생에 관한 연구

김상근¹, 이만휘²

¹충남대 수의대
²

Abstract

This study was carried out to investigate the optimal activation condition for parthenogenetic development. In order to activate oocytes at 24 hrs post onset of maturation, the oocytes were cultured $3{sim}13{mu}M;Ca^{2+}$ for 5 min., $5-8{mu}g/ml$ cytoclacin for 6 hrs, 0.5~2.0 mM 6-dimethylaminopurine(DMAP) for 3 hrs alone or combination. The activated oocytes were cultured in TCM-199 media at 5% $CO_2$, 95% air, $38^{circ}C$. The cleavage rate after 48 hrs culture of oocytes treated with $3-13{mu}M;Ca^{2+}$, $5-8{mu}g/ml$ cytoclacin and 0.5~2.0 mM DMAP for 5 min., 6 hrs and 3 hrs were 9.6%~20.0%, 0.0%~7.3% and 9.4%~21.8%, 0.0%~7.3% and 9.1%~21.8% and 0.0%~7.3%, respectively. When oocyte were treated with $10{mu}M;Ca^{2+}$, $10{mu}g/ml$ cytoclacin and 2.0 mM DMAP the blastocyst formation rate was significantly higher than other group. The cleavage rate after 48 hrs culture of oocytes treated with $Ca^{2+}$ + cytoclacin, $Ca^{2+}$ + DMAP, cytoclacin + DMAP were 75.9%~93.5% and 9.7%~19.0%, respectively. When oocytes were treated with $Ca^{2+}$ followed by DMAP, the blastocyst formation rate was significantly higher than other group(p<0.05). When necleus transferred embryos co-cultured with bovine serum albumin(BSA), epithemal growth factor(EGF) and calf serum(CS), the developmental rate to blastocyst were higher than control group.

Key Words: parthenogenesis, activation, $Ca^{2+}$, cytoclacin, DMAP