维生素C对藏西北绒山羊精液冷藏效果及其全基因组DNA甲基化的影响

【目的】研究维生素C对4℃保存藏西北绒山羊精液效果及其全基因组DNA甲基化的影响,为高效利用藏西北绒山羊种公羊精液及推广其人工授精技术打下基础。【方法】假阴道人工采集藏西北绒山羊种公羊新鲜精液,用维生素B12进行4倍稀释后添加不同浓度的维生素C(0、2.0、4.0、6.0、8.0、10.0和12.0 mg/m L),4℃保存24 h,观察统计其精子活力、精子顶体完整率和质膜完整率,并通过实时荧光定量PCR检测精子全基因组DNA甲基化的情况。【结果】藏西北绒山羊精液4℃保存24 h后,以添加8.0 mg/m L维生素C的效果最佳,对应的精子活力、精子顶体完整率和质膜完整率分别为(73.41±0.47)%、(58.89±0.88)%和(68.76±0.81)%,3个指标均极显著高于未添加维生素C的空白对照组(P0.01)。添加8.0 mg/m L维生素C的藏西北绒山羊精子全基因组DNA甲基化水平与新鲜精液的接近,均显著低于未添加维生素C的冷藏精液(P0.05)。【结论】原精液中添加2.0 mg/m L维生素C可降低4℃保存藏西北绒山羊精液的全基因组DNA甲基化异常水平,提高藏西北绒山羊精液冷藏效果。

Effects of vitamin C on Northwest Tibet Cashmere Goat semen preserved at 4℃and levels of complete genome DNA methylation

Objective]This study investigated the effects of vitamin C on Northwest Tibet Cashmere Goat semen pre-served at 4℃and levels of the complete genome DNA methylation, in order to lay a foundation for utilization of ram semen and promotion of artificial insemination technique in the future . Method]Fresh semen of ram of Northwest Tibet Cashmere Goat was collected using artificial Vagina. Vitamin C of different concentrations(0,2.0,4.0,6.0,8.0,10.0 and 12.0 mg/mL) were added into the semen 4-time diluted with vitamin B12 and preserved at 4℃for 24 hours. Sperm motili-ty, acrosomal integrity and plasma membrabe integrity were observed and complete genome DNA methylation was detected by real-time fluorescence quantitative PCR. Result]The Northwest Tibet Cashmere Goat semen after 24 hours preserved at 4℃with 8.0 mg/mL vitamin C showed the best quality. Sperm motility was(73.41±0.47)%, acrosomal integrity(58.89± 0.88)% and plasma membrabe integrity(68.76±0.81)%. The three indexes were higher than those of control which added no vitamin C(P<0.01). At the same time, the level of sperm complete genome DNA methylation was close to that of fresh semen, and significantly lower than those of semen added no vitamin C(P<0.05). Conclusion]Adding 2.0 mg/mL vitamin C in Northwest Tibet Cashmere Goat semen preserved at 4 ℃ can reduce the level of DNA methylation and increase the quality of sperm preserved at refrigeration temperature.