| Mammalian somatic nuclear transplantation is a complicated and orderful process.This paper researched the processes of sheep oocyte in vitro maturation,parthenogetic and the reconstruction of embryo,and established a series of more steady nuclear transfer system.The results are as follows: 1. The influencing factors concern sheep oocyte IVM in culture medium were studied though 3 methods . That was:the concentration of epithelial growth factor(EGF), the concentration of FSH and LH and the addition of different diameter follicule liquid. It showed that(:1)30 ng/ml EGF could not only inhance the maturation of sheep oocytes (85.0% vs 60.0%, 71.7%, 72.7%, P﹤0.05), but was useful for sheep parthenogentic embryo development;(2)FSH 1.0μg/ml,LH　5.0IU/ml　was　proper　concentration　for　sheep　oocyte　IVM;(3)Addition of 3% sheep big follicle liquid to OM was significantly higher than the team of addition of 20% sheep small follicle liquid in the rate of oocytes maturation( 78.6% vs 36.5%, P﹤0.01 ) ,and was higher than addition of 3% sheep small follicle liquid and the control(78.6% vs60.0%, 67.5%, P<0.05). 2. This studay was to compare the effects of differrent development stage of oocyte on the development capacity of OPS vitrifying sheep oocytes. The results showed:all groups of differrent development stage of oocyte had no differrence on the rate of morphologically normal,24h group had a higher rate of cleavage and blastocyst than other groups(38.3% vs 13.3%,26.7%,28.3%;13.0% vs 0%,6.3%,5.9%, P < 0.05),but there had no differrence between 18h group and 26h group(P < 0.05).It showed that24h group could gain the best effects of OPS vitrification. 3. The cleavage and development of sheep oocytes activated by different chemical activative procedures were investigated. There was no significant difference in morula/blastocyst development between ionomycin-6-DMAP and ionomycin 6-DMAP+CB sequential treatments ,but the two treatments had a higher rate of morula/blastocyst development( P < 0.05) compared to ionomycin-CB treatment and were significantly higher than only ionomycin treatment(P <0.01); ethanol + 6-DMAP+CB had a higher rate of blastocyst development than ethanol-6-DMAP and ethanol-CB(P <0.05),and was significantly higher than only ethanol treatment(P <0.01). 4. Effects of parameters of electrical activation on the development of sheep parthenogentic embryo was studied.Comparing with 0.8kv/cm,2.0kv/cm,1.2kv/cm,1.6kv/cm have the better activation rate(P <0.05);3 pulses has the best activation efficiency(P <0.01) . 5. These examinations analysised the impacts of the age of oocyte and the oocyte cycle stage (MII and TII ) on sheep nuclear transfer.The results showed :(1)The appropriate enucleation time is 18~21h for sheep nuclear transfer; (2) TII stage method could enhance the enucleation efficiency significantly(P < 0.05),but the rate of morula/blastocyst that obtained from both methods had no significant difference(P > 0.05). 6. Effects of oocyte'hyperosomolarity treatment and donor cell starvation on sheep NT were explored. Hyperosomolaried treatment could significantly improve the efficiency of inculeation , but had no effects on the development of NT embryo ; Granulosa serum starvation method and high-confluncy method had no difference ,but the two methods were higher than control group at the fusion rate(P < 0.05).
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