“两步法”体外培养山羊卵母细胞的初步研究  

Two-step" Protocol for Oocytes Maturation in Vitro

作　　者：邵华[1] 孟和宝力高 张锁链[1] 旭日干[1] 

机构地区：[1]内蒙古大学实验动物研究中心,内蒙古呼和浩特010021

出　　处：《内蒙古大学学报（自然科学版）》2002年第5期546-551,共6页Journal of Inner Mongolia University：Natural Science Edition

基　　金：国家自然科学基金资助项目 (批准号 3 9860 0 5 6)

摘　　要：体外成熟培养的卵母细胞经体外受精后 ,囊胚发育率远低于体内成熟卵母细胞 .原因之一是由于体外成熟卵母细胞的不充分获能造成的 .有假说认为 :卵母细胞在体外培养过程中 ,如果延长 GV期 ,可促进卵母细胞进一步获能 ,从而提高发育潜能 .本文以山羊卵子为研究对象 ,探讨了山羊卵泡各种成分以及异种家畜卵泡液对山羊卵母细胞核成熟抑制以及抑制解除后减数分裂的影响 .结果显示 ,山羊 5 0 % Q卵泡液、1 0 0 %卵泡液、半卵泡、打孔整体卵泡以及完整卵泡对山羊卵母细胞的核成熟抑制率分别为 :0 .0 0 %、9.3 8%、60 .47%、78.2 6%和70 .0 9% .随后 ,对经山羊半卵泡、打孔整体卵泡、完整卵泡抑制培养后的卵子进行恢复培养发现 ,卵子核成熟率分别为 :64.2 9%、3 .49%和 4.1 3 % .山羊卵母细胞与半卵泡共培养 2 4h后进行 0 h、1 2 h、2 0 h、2 4h和 3 0 h的恢复培养 ,核成率分别为 :0 .0 0 %、2 0 .0 0 %、61 .90 %、64.2 9%和 43 .75 % .表明 ,山羊半卵泡可以有效抑制山羊卵母细胞使之处于 GV期 ,并且这种抑制作用可以恢复 ,恢复培养时间以 2 0 h和 2 4h较为适宜 .初步研究异种家畜卵泡液对山羊卵母细胞核成熟抑制的影响发现 ,5 0 %牛卵泡液、1 0 0 %牛卵泡液、5 0 %绵羊卵泡液、1 0 0 %绵羊卵泡液对山羊卵母细?The developmental frequency of blastocyst stage embryos derived from cumulus oocyte complexes (COCs) matured,fertilized and cultured in vitro is inferior to that derived from COCs matured in vivo.One of the reasons is that oocytes matured in vitro have been not acquired full developmental competence,and their devlopmental potential is decreased.Therefore,it is hypothesized that if oocyte can be cultured in vitro under condition that maintains meiotic arrest at the germinal vescle (GV) stage,then they may have the opportunity to acquire greater developmental competence.In this study,goat oocytes were cultured in different methods,including addition of follicular fluid (FF) to the culture medium,culturing within follicle hemisections,hole pierced intact antral follicle and intact antral follicle,so that physiological methods can be established for the maintainence of meiotic arrest.The results show:frequencies of meiotic inhibition on oocytes in 50% gFF,100g FF,follicle hemisection,hole pierced intact antral follicle and intact antral follicle,were 0.00%,9.38%,60.47%,78.26%,70.09% respectively (a,b P<0.01).Oocytes were cultured in maturation medium for 24 h after inhibited in fllicle hemisection,hole pierced intact antral follicle and intact antral follicle for 24 h respectively,the frequencies of matured oocytes were 64.29%, 3.49% and 4.13% respectively (c,d,P<0.01).Before and after maturation culture with hole pierced intact antral follicle,the frequencies of inhibition were 78.26%,61.63% respectively.Follicle hemisection is the best meiotic arrest factor.Inhibition frequency of oocytes cultured with follicle hemisection were decreased step by step following co culture time prolonged.Goat oocytes inhibited by follicle hemisection for 24 h were matured in vitro for 0h,12h,20h,24h and 30h,the frequencies of maturation were 0.00%,20.00%,61.90%.64.29% and 43.75% respectively.The maturation frequencies at 20h,24 h and 30h showed no significant difference,but the frequency of abnormal oocytes at 30h increased

关 键 词：体外培养 山羊 卵母细胞 核成熟抑制 恢复培养 两步法 体外成熟 细胞培养 

分 类 号：S827.3[农业科学—畜牧学] Q813.7[农业科学—畜牧兽医]