| Many influence factors within IVF were experimented systematically in this study for the sake of establishing an acceptable IVF system on cow and calf. The main contents of this study are: effect of different factors on cow and calf IVF technique.The oocytes were collected from slaughtered cow ovaries. The influence of 10% BFF supplementation of IVM media on oocytes maturation and embryo development, effect of centrifugal speed on IVF, effect of different culture method on embryo development and effect of different culture condition on embryo development had been studied. The results as follow: The first polar body extruding,cleavage rate and blastula rate between the group of supplied 10%FBS and the group of supplied 10%FBS and 10%BFF were not significantly different (80.0% vs 76.7%;55.8% vs 56.3%;20.69% vs 21.4). But the embryo development ability in the latter group were better than the embryos in the former group. The cleavage rates of embryos centrifuging at 750 rpm and 1000 rpm were significantly higher than the embryos centrifuging at 1500 rpm (P<0.05). and the cleavage rate in the 1000 rpm group was the highest, but the difference was not significantly when compared to the group of 750 rpm (P>0.05) . the blastula rate in the groups of tow-step method and three-step method were not significantly different(28.2% vs 28.9%, P>0.05), but the difference of cleavage rates was significant(59.2% vs 50.0%, P<0.05). Embryos cultured in 5%CO2 in air, co-cultured with cumulus cells or in 5% CO2, 5% O2, and 90% N2, the cleavage rates in the three groups were not significantly different(44.3%,50.8% and 48.4%, P>0.05), and the blastula rates in the latter two groups were also not significantly different (32.3% vs 30.0%,P>0.05), but significantly higher than the rate in the first group(P<0.05). The mammalian blastocyst comprises an inner cell mass (ICM) and a trophectoderm cell layer(TE). The TE of cultured bovine blastocysts was stained when blastocysts were treated with a permeabilizing solution containing the ionic detergent Triton X-100 and the fluorochrome propidium iodide. The result show that 60~70s was the best time when blastocyst conducted by Triton X-100. And using this methord ,we test the effect of fructose on the embryo in vitro development of cow. The data show that compared with the glucose group, the number of ICM and TE are both higher(43.6 vs 40.2, 99.3 vs 89.9), but not significantly (P>0.05). The study indicates that, fructose up to 1.5mM concentration can be used as an alternative for energy substrate in culture media. In the study of Holstein juvenile calf superovulation and in vitro fertilization, 10 juvenile calves ,aged 4~9w, were subjected to gonadotropin stimulation to evaluate the effect of age, culture condition and individual donor variation on JIVET(Juvenile In Vitro Embryo Transfer). 31.8±9.75(mean±S.D.) cumulus–oocyte complexes (COCs) per donor were recovered from group 1 aged 30~40d and 21.8±11.67 COCs from group 2 aged 46~63d.The differences were not significant. No differences were observed in cleavage rates (48.5% vs 54.1%) between low O2(5% CO2, 5% O2 ,90% N2) and co-cultured(20%O2, 95% air) conditions after in vitro fertilization(IVF). However, the morulae rate and were very significantly different(0% vs 15.2%). There was a significant variation between individual donors. Overall, 70% (7/10) of the calves exhibited follicle development.
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