提高牛体外性控胚胎效率关键技术研究

为了提高牛体外受精效果,试验从性控冷冻精液体外获能方法及抗氧化剂的添加、体外成熟24h后卵母细胞是否部分脱除卵丘细胞3个方面,研究了影响牛性控精液体外受精效率的关键技术。结果表明,一次离心体外获能法体外受精卵裂率[(68.3±3.7)%]、囊胚率[(30.9±2.3)%]与精子上浮获能法体外受精卵裂率[(67.8±2.6)%]、囊胚率[(29.7±3.7)%]无显著差异(P>0.05),但显著高于二次离心体外获能法体外受精卵裂率[(56.1±4.1)%]、囊胚率[(21.6±4.6)%](P<0.05);获能液和体外受精液添加1.0%抗坏血酸组体外受精卵裂率[(70.2±3.2)%]、囊胚率[(35.0±4.7)%]显著高于不添加组体外受精卵裂率[(60.0±4.5)%]、囊胚率[(28.3±3.6)%](P<0.05);在体外受精前,成熟卵母细胞周围的卵丘细胞脱除情况为部分脱除组体外受精卵裂率[(70.1±4.6)%]、囊胚率[(35.5±3.7)%]显著高于不脱除组体外受精卵裂率[(52.9±4.1)%]、囊胚率[(26.1±3.6)%](P<0.05)。由此可见,性控精液体外受精时,获能方法宜采用一次离心体外获能法或上浮获能法,且精子获能液和体外受精液中宜添加抗坏血酸;体外受精前,成熟卵母细胞部分脱除卵丘细胞对于提高性控精液体外受精胚胎发育率具有重要意义。

Development of Key Technology for Improving in Vitro Fertilization Efficacy of Bovine Sexing Embryo

In an attempt to improve the in vitro fertilization(IVF)efficacy of bovine sexing embryo,a set of key technology for improving IVF efficacy of bovine sexing semen was developed.Firstly,the effect of three in vitro capacitation methods of bovine sexing frozen semen and antioxidant additive addition on IVF efficacy of bovine sexing semen was statistically evaluated;secondly,the impact of partial removal of cumulus cell from oocyte matured in vitro for 24 h on IVF efficacy of bovine sexing embryo was assessed.The results showed that while the cleavage rate and blastocyst rate of IVF produced zygotes of one-time centrifugation in vitro capacitation method[(68.3±3.7)%,(30.9±2.3)%]had no significant differences with sperm floating capacitation method[(67.8±2.6)%,(29.7±3.7)%](P>0.05),they were significantly higher than those of two-time centrifugation in vitro capacitation method[(56.1±4.1)%,(21.6±4.6)%](P<0.05);the treatment with addition of 1.0%ascorbic acid in capacitation liquid and IVF liquid had significantly higher cleavage rate[(70.2±3.2)%]and blastocyst rate[(35.0±4.7)%]of IVF produced zygotes compared to the treatment without additive[(60.0±4.5)%,(28.3±3.6)%](P<0.05);the cleavage rate and blastocyst rate of IVF produced zygotes in the treatment that the surrounding cumulus cell was partially removed from the oocyte prior to IVF[(70.1±4.6)%,(35.5±3.7)%]were found significantly higher compared to the treatment without cumulus cell removal[(52.9±4.1)%,(26.1±3.6)%](P<0.05).The combined data suggest that the one-time centrifugation in vitro capacitation method and sperm floating capacitation method have better performance in improving IVF efficacy of bovine sexing semen,and the addition of 1.0%ascorbic acid in capacitation liquid and IVF liquid is recommended.In addition,the partial removal of cumulus cell from oocyte prior to IVF has important significance on improving the development of IVF embryo produced by sexing semen.