Study On Buffalo-bovine Interspecies Somatic Cell Nuclear Transfer

Interspecies somatic cell nuclear transfer (NT) may be used as an effective method to conserve endangered species and to support the study of nuclear-cytoplasm interaction. This experiment adopted somatic cell nuclear transfer technologies to research on the buffalo-bovine interspecies NT. The maturation time of recipient cytoplasm, treatment of donor cells, parthenogenetic activation of recipient oocyte, and embryo culture medium were mainly researched; the interspecies NT technologies in our laboratory was established, and the efficiency of buffalo-bovine interspecies NT was compared with that of buffalo and bovine intraspecies NT.1 Somatic cell nuclear transfer:1) Firstly, the parthenogenetic activation of bovine oocytes with chemical simulation was investigated. The activation rate of the complex chemical activation was higher than that of the single chemical one, and the complex of lonomycin+6-DMAP got the best result (8-cell rate 22.0%).2) Secondly, we compared the development of the bovine parthenogenetic embryos cultured in TCM199 and CR1aa and found bovine embryos was suitable to be cultured in CRlaa (21.7% vs. 16.7%, p<0.05).3) Effects of in vitro maturation (IVM) duration on maturation and enucleation rate of bovine oocytes were studied. The maturation rate of oocytes matured for IVM 16-18h was significantly lower than that of IVM for 19~21h (45.6% vs. 60.8%, p<0.05) and 22-24h(45.6% vs. 64.7%, p<0.05). The enucleation rate of oocytes matured for IVM 22~24h was significantly lower than that of IVM for 16~18h (52.2% vs. 80.6%, p<0.05) and 19~21h (52.2% vs. 73.3%, p<0.05). The results indicate that maturation rate of bovine oocytes increase as the IVM duration prolongs, while the enucleation rate decreases. To advance the enucleation rate on NT, we selected the oocytes that IVM 20h as cytoplasmic recipients.