| 摘要: |
| 旨在探讨提高猪GV期卵母细胞冷冻保存效率的可能途径。将EDS、EFS40和ES冷冻保护液用作卵母细胞冻前、冻后程序的处理,但不冷冻,比较3种冷冻保护剂对猪GV期卵母细胞的毒性作用;采用ES液作玻璃化液,比较常规细管法、OPS法和电镜铜网法3种冷冻载体对猪GV期卵母细胞的冷冻效果;并以ES液作玻璃化液,OPS管为冷冻载体,将猪GV期卵母细胞分5组,即对照组、CB＋离心处理组、直接冷冻组、CB＋冷冻组、CB＋离心＋冷冻组进行对比处理,比较各处理卵母细胞的冻后存活率与发育率。结果表明,在不同冷冻保护液中,以ES液组合作为玻璃化冷冻液时的毒性作用最低,与对照组间无明显差异（P＞0.05）;在3种冷冻载体中,用OPS法冷冻猪GV期卵母细胞,所获冻后存活率明显高于电镜铜网法和细管法（65.4%对45.0%和38.6%,P＜0.05）,并可获得最佳的冻后成熟率（43.3%）;猪GV期卵母细胞单纯经细胞松弛素B和离心极化处理,不会严重影响卵母细胞的存活,但卵裂率显著低于对照组（39.0%对52.1%,P＜0.05）;细胞松弛素B处理或细胞松弛素B＋离心极化处理后再进行玻璃化冷冻,并不能提高卵母细胞的冻后存活率与发育率。采用OPS法直接进行GV期卵母细胞的玻璃化冷冻,可获得7.8%的冻后卵裂率,并能获得桑椹胚发育,是一种有效的猪卵母细胞冷冻保存技术。 |
| 关键词: 猪 卵母细胞 玻璃化冷冻 |
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| 基金项目:上海市科技兴农重点攻关项目（沪农科攻字［2003］第14-1号）；国家自然科学基金项目（30270958） |
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| Study on vitrification of porcine GV stage oocytes |
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| Abstract: |
| In order to investigate an approach to improve the cryopreservation efficiency of porcine GV stage oocytes,three kinds of cryoprotectants(EDS,EFS40 and ES) were respectively used for the treatment of pre-and post-vitrification of oocytes.Then,with the ES chosen for vitrification solution,the routine straw method,OPS method and electron microscopy grid(EMG) method were adopted to cryopreserve porcine GV stage oocytes.Finally,ES was used as vitrification solution and OPS method as vitrification carrier,and porcine GV stage oocytes were divided into 5 groups for the treatment: the control,CB+ centrifugation group,the group of vitrification only,CB+ vitrification group,and CB+ centrifugation+vitrification group.The toxic experiment of different cryoprotectants on porcine GV stage oocytes showed that ES solution showed the lowest toxicity and there was no significant difference compared with the control(P>0.05).Among 3 types of freezing carriers,OPS method yielded significantly higher post-warming survival rate than EMG and straw method did(65.4% vs 45.0% and 38.6%,P<0.05),and resulted in the optimal post-warming maturation rate(43.3%) as well.Porcine GV stage oocytes treated with cytochalasin B and centrifugation separately didn't show significant influence on decrease or increase in survival.However,their cleavage rate would be significantly lower than that of the control(39.2% vs 52.1%,P<0.05).The pre-treatment with cytochalasin B or cytochalasin B+centrifugation could not improve the post-warming survival and developmental rates of vitrified GV stage oocytes.The OPS vitrification of porcine GV stage oocytes without other treatments could result in 7.8% of cleavage rate and subsequent morula development. |
| Key words: pig oocytes vitrification |
