本文關(guān)鍵詞：驢卵母細(xì)胞成熟及GV期玻璃化冷凍的研究　出處：《內(nèi)蒙古農(nóng)業(yè)大學(xué)》2016年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 驢 GV期卵母細(xì)胞 玻璃化冷凍 微絲 線粒體

【摘要】：本實(shí)驗(yàn)主要研究不同采集方法回收卵母細(xì)胞的效率及不同濃度的FSH對卵母細(xì)胞成熟率的影響,同時(shí)對GV期的卵母細(xì)胞進(jìn)行玻璃化冷凍,并對其顯微結(jié)構(gòu)進(jìn)行觀察,研究結(jié)果如下：1.切割法獲得卵母細(xì)胞成功率為81.57%,綜合法獲得卵母細(xì)胞成功率為79.01%,兩者差異不顯著(P0.05),而利用抽吸法獲得卵母細(xì)胞成功率為48.09%,顯著低于抽吸法和綜合法(P0.05)。2.卵母細(xì)胞成熟培養(yǎng)體系中分別添加不同濃度的FSH,根據(jù)成熟率的不同來分析FSH的最適濃度,結(jié)果表明60μg/mL、80μg/mL的FSH的成熟率(21.43%、38.98%)顯著低于添加100μg/mL、120μg/mLFSH的成熟率(51.77%、47.29%)(P0.05),添加100μg/mLFSH成熟體系的成熟率略高于120μg/mL的成熟體系,但差異不顯著(P0.05)。3.本試驗(yàn)比較了不同培養(yǎng)時(shí)間對卵母細(xì)胞成熟率的影響,結(jié)果表明隨著培養(yǎng)時(shí)間的延長,成熟率穩(wěn)步提高,到達(dá)30～32 h后成熟率趨于穩(wěn)定。4.采用四個(gè)濃度梯度(Ⅰ、Ⅱ、Ⅲ、Ⅳ)分別對GV期卵母細(xì)胞進(jìn)行玻璃化冷凍,解凍后體外成熟對其形態(tài)正常率和成熟率進(jìn)行統(tǒng)計(jì),結(jié)果表明四種冷凍保護(hù)劑成熟培養(yǎng)后卵母細(xì)胞的形態(tài)正常率分別為84.62%、87.50%、71.43%、68.63%,差異不顯著(P0.05),其中冷凍液Ⅱ的成熟率(28.13%)顯著高于其他三組的成熟率,但都顯著低于對照組。由此可知最佳的冷凍劑為II組：20%DMSO+20%EG+60%M199+0.25M Sucrose。5.分別選用普通玻璃細(xì)管、OPS管、GMP管作為冷凍載體冷凍GV期驢卵母細(xì)胞,解凍后成熟培養(yǎng)并對其形態(tài)正常率和成熟率進(jìn)行統(tǒng)計(jì),發(fā)現(xiàn)卵母細(xì)胞形態(tài)正常率分別為81.67%、87.50%、76.47%,差異不顯著(P0.05)；OPS管組卵母細(xì)胞成熟率(28.13%)顯著高于普通玻璃細(xì)管組(11.67%),但與GMP管組(23.53%)差異不顯著,因此OPS管是最合適的冷凍載體。6.本試驗(yàn)還對正常卵母細(xì)胞及解凍后的卵母細(xì)胞進(jìn)行微絲及線粒體的免疫熒光染色,免疫熒光結(jié)果顯示冷凍組卵母細(xì)胞微絲在皮質(zhì)區(qū)的分布明顯減少,部分微絲雜亂分布于胞質(zhì)中央,冷凍組卵母細(xì)胞的線粒體數(shù)量明顯低于正常組,由此可以說明冷凍對卵母細(xì)胞微觀結(jié)構(gòu)有損傷。
[Abstract]:The effect of different collection methods on oocyte recovery efficiency and FSH concentration on the maturation rate of oocytes was studied. At the same time, vitrification of GV phase oocytes was carried out. The results were as follows: 1. The success rate of oocytes obtained by cutting method was 81.57, and the success rate of oocytes obtained by comprehensive method was 79.01%. There was no significant difference between the two groups (P 0.05), but the success rate of oocytes obtained by aspiration was 48.09%. It was significantly lower than that of suction method and comprehensive method (P0.05 路2.The optimum concentration of FSH was analyzed according to the different maturation rate of FSH). The results showed that the maturation rate of 60 渭 g / mL 80 渭 g / mL FSH was significantly lower than that of adding 100 渭 g / mL. The maturation rate of 120 渭 g / mLFSH was 51.77 and 47.29, respectively. The maturation rate of the matured system added 100 渭 g / mLFSH was slightly higher than that of 120 渭 g / mL maturation system. The effect of different culture time on the maturation rate of oocytes was compared. The results showed that the maturation rate increased steadily with the extension of culture time. The maturation rate of GV oocytes was stable at 30 h. 4. Four concentration gradients (鈪，

本文編號：1387992