Study On In Vitro Maturation, Parthenogenetic Activation And Somatic Cell Nuclear Transfer In Sheep

This study was set out to investigate the superovulation effect of hormone from different manufacturers on young lamb, to compare the ablity of COCs in vitro maturation, oocyte parthenogenetic activation, cloned embryos development between young lamb and adult sheep, as an attempt to establish a steady nuclear transfer system. Results of this study mainly include the following:1.To discourse the effect of different medicines(Ningbo,CAS,Canada)on young lamb superovulation. The percentage of useable oocytes and average number of useable oocytes were(69.6% vs 87.9% vs 88.4%, P<0.05)and(15.1 vs 25.1 vs 27.1, P<0.05), the percentage of useable oocytes and average number of useable oocytes with Ningbo were significantly lower than the other two gonadotropins, it showed that Ningbo hormone was inferior to the other two hormone, however, the result of Canada was higher 0.5% and 2 than CAS, respectively. It turns out that Canada hormone was the best.2. The maturation rate of adult sheep with the FBS in vitro maturation culture was higher(77.3%)than the maturation rate of young lamb(51.1%)(P<0.05),and the maturation rate of adult sheep with the ESS in vitro maturation culture was higher(78.9%)than the maturation rate of young lamb(59.0%)(P<0.05), it showed that the effect of adult sheep oocyte was much better than the young lamb oocyte. But the maturation rates between the two young lamb groups were significantly different(P<0.05), the maturation rate with the ESS in vitro maturation culture was 7.9% higher than that with the FBS, it showed that the ESS can enhance the maturation rate in vitro maturation of the young lamb.3. Two actuvating agents of ionomycin with 6-DMAP, A23187 with 6-DMAP were compared, the cleavage rates and blastocyst rates of two groups of young lamb were(50.7% vs 48.0%, P<0.05) and(13.4% vs 11.1%, P<0.05), the cleavage rates and blastocyst rates of two groups of adult sheep were(69.5% vs 67.6%, P<0.05)and(29.5% vs 28.6%, P<0.05), respectly, there was no significant difference. But the cleavage rate and blastocyst rate of the first young lamb were 2.7%and 2.3% higher than the second group, and the cleavage rate and blastocyst rate of the first adult sheep were 1.9%and 0.9% higher than the second group, however, the cleavage rates and blastocyst rates of adult sheep were significantly higher than the young lamb(P>0.05). it showed that both of the activation methods could activate the sheep oocyte effectively, but the development of adult sheep activated oocyte was better than the young lamb oocyte.4. Comparing the enucleated effect of the young lamb and adult sheep oocyte after 22~24h in vitro maturation, the number of enucleated oocyte and enucleated rate were(72 vs 75)and(60.0% vs 50.3%, P>0.05), the enucleated rate of the young lamb oocyte was 9.7% higher than the adult sheep, the difference was significant(P>0.05), it showed that the enucleated effect of the young lamb oocyte was better than adult sheep oocyte after 22~24h in vitro maturation.5. The cleavage rates and blastocyst rates of the young lamb and adult sheep NT embryo were(29.2% vs 58.9%, P>0.05)and(5.0% vs 17.0%, P>0.05), the cleavage rate and blastocyst rate of adult sheep were 29.8% and 12% higher than the young lamb, respectively. The difference was significant(P>0.05). It showed that the development of the young lamb NT embryo was worse than adult sheep NT embryo.