Study On Main Influences Of The Effect Of Intracytoplasmic Sperm Injection(ICSI) In Goat

The study was conducted to improve the total impact of Intracytoplasmic Sperm Injection (ICSI), by investigating main influences factors to ICSI. In the ICSI process, test method was modified so that we can reduce oocyte damage bringing by mechanical, biological and reagent stimulation, and we could finally get a improved sperm nucleus decondensation rate (fertilization rate).When dealed sperm in media with 8%(g/ml)PVP content, we got a comparative higher operating rate compared with dealing in 5%PVP media (92.50%vs44.62%, P<0.01), comparing with dealing sperm in 5%PVP media, a comparative higher fertilization (58.33% vs 29.73%, P<0. 05) also be founded out. By using 8μm caliber injection pipe, we got a comparative low sperm decondensation while comparative higher injection succeeding rate in an action and fertilization rate, so the writer considered injection pipe of 8μm caliber as the just choice for ICSI in goat. Doing ICSI with goat oocyte matured 26h or 28h in vitro, the cleavage rate, mulberry rate and blastula rate were all remarkable (or significantly) higher than mature 24h group, so we thought, that prolong in vitro mature time properly do favor in enhancing ICSI effects in goat. Treating sperm in the media supplemented with 40μg heparin enhanced fertilization rate from 46.67% to 57.17%, and a 10.5 percent increasing extend, it was obvious that the test result benefited from heparin. As the ICSI result was concerned, the cleavage rate of the oocyte injected with sperm treated by cytochalasin B (CB) was not good than that of the untreated ones, nor the mulberry rate between the two groups had significant differences. The ICSI fertilization rate of frozen semen was significantly higher than that of fresh semen (52.21% vs 26.44%, P<0.05). We enactmented three groups to study on oocyte activating method, control group (dealed with no activation after injection), test group One(co-activated oocyte after injection, with 'Ion 5min+6-DMAP 3h' ) and test group Two(activated oocyte after injection, with single 'Ion 15min'), and the result was, oocytes in control group had been hardly activated (with 7.94% cleavage rate only, and getting no mulberry), the cleavge rate, mulberry rate and blastula rate of test group One and Two were, 82.69% & 78.40%, 47.67% & 46.94%, and 13.95% & 9.18% respectively, all the data were significantly higher than the same data in control group, while there were no significantly differences in data between the two test groups. We did a research on the influences of cumulus cells and co-culturing with single cumulus cell to the early development of goat ICSI embryo, there were there test groups as well, group One (the oocytes were bare, without any co-culturing after ICSI ), group Two (the oocytes were bare, and were co-cultured after ICSI ) and group Three (there were some cumulus cells on the oocyte, without any coculturing). We found that, the cleavage rates of the three groups have not gotten significantly differences(P>0.05), but Group Two and Group Three were all significantly higher than Group One in mulberry rate(44.86% vs 29.09% and 46.77% vs 29.09%, P<0.05) and in blastula rate(19.30% vs 7.27% and 20.97% vs 7.27%, P<0.05). There were significantly differences (P>0.05) between cleavage rate and mulberry rate of ICSI embryo in high oxygen condition (atmosphere with 20%O₂) and low oxygen gas condition (5%CO₂+5%O₂+N₂90%), while the blastula rate was significantly higher than the corresponding datum in low oxygen gas condition(19.59%vs7.81 %, P<0.05).