Research On In Vitro Embryo Production In Sheep

It is well known that artificial insemination (AI). embryo transfer (ET) and in vitro fertilization (IVF) are among the most efficient techniques. While IVF technology in sheep is still unskilled,and it need more study before practical use.Ovaries obtained from the slaughterhouse and matured in vitro were used to reseach the factors which reported influence the results of IVF and early embryo culture in order to optimized the technology of IVF in sheep. The major results obtained are as follows:1 The recovery of COCs from slaughtered sheep ovary by surface — follicles cutting is efficient compared with the aspiration and more COCs could be recovered by surface-follicles cutting (4.47per ovary vs 2.51 per ovary). But the time used is a little longger(55s per ovary vs51s per ovary)2 Addition with different dose of FSH can stimulate the maturation of oocytes in vitro and the percent of maturation did not differ between 1 μ g/mk 5 μ g/ml and 10 μ g/ml.However both the 1 μg/ml and 5 μ g/ml treatments result in cleavage rate increase compared with 10 μ g/ml treatments3 Oocytes was incubated in maturation media with addition of EGF as both the 50 ng/ml and 100 ng/ml treatments displayed a significant increase (P < 0.05) in the maturation rate , but only in the 100 ng/ml EGF treatment, which displayed a significant (P < 0.05) increase in cleavage rate.4 Oocytes were co-cultured with granulosa cells of certain concentration ( 2-6×10~6 /ml)had no obvious effect on IVM compared with oocytes co-cultured without granulosa cells, but the former method can greatly increased the percentage of cleavage ( 55.6% vs 53.8% ) and morulae/blastocyst.(28.9% vs 19.1%) (P<0.05).5 The effect of individual rams cannot be neglect. The result of our experiment showed that when sperms coming from three different rams treated with the same method and used for IVF /There was significant difference in rates of cleavage ( 54. 7%, 41.3%, 43.3%) and morulae/blastocyst (28.2%, 19. 7%, 22. 6%) among different individual rams(P<0. 05).6 When oocytes and sperm were incubated for 8h, 12h, 16h 20h respectively. The highest rates of cleavage and morulae/blastocysts were obtained from 16h.7 When sheep' embryos were co-cultured with different meduim I: TCM199+GCM+15%ESS+EGF ; II: TCM199 +15%ESS+EGF; III: SOF fluid+15%ESS+10 μg/ml NEAA+20 μ g/mlEAA+ EGF; IV: SOFfluid+BSA+10 μ g/mlNEAA+20 μ g/mlEAA+ EGF TCM199, The result showed that the cleavage rates of team Ⅰ,Ⅱ,Ⅲ was significant higher than team Ⅳ(P < 0.05),and team I was higher than team Ⅱ ,result of team Ⅲ resembled Ⅰ and Ⅱ; The results also showed that team I got the higher percentage of morulae/blastocysts ,but there is no significant difference when compared with other mediums8 The Gradual Changed TCM199 Culture System for IVF on Sheep can significant enhanced the rates of cleavage and morulae/blastocysts compared with the regular method (P < 0.01).This is an effective way to progress sheep's Embryo in vitro fertilization technology .