核移植方法和维生素C影响绵羊克隆胚胎体外发育

doi:10.11843/j.issn.0366-6964.2014.09.005

摘要/Abstract

摘要：

旨在研究绵羊体细胞核移植过程中，不同核移植方法及胚胎培养液中添加维生素C对绵羊克隆胚胎生产效率的影响，为优化绵羊体细胞克隆技术体系提供试验依据。用屠宰场采集的绵羊卵巢为试验材料，经卵母细胞体外成熟后进行核移植操作，采用微电极融合、无透明带克隆与显微注射等核移植方法，比较胚胎构建效率，利用绵羊孤雌胚胎摸索发育液中添加维生素C最适浓度，将该浓度用于克隆胚胎体外培养，统计胚胎发育情况。结果表明:采用25 V微电极融合法融合率显著高于平行电极融合法（P<0.05），但与30 V电压差异不显著。25 V电压微电极融合法死卵率显著低于30 V融合法（P<0.05）；无透明带构建克隆胚胎的桑椹胚率显著高于显微注射法（P<0.05）,但囊胚率差异不显著（P>0.05）；胚胎培养液中添加50 μmol•L^-1维生素C显著提高了孤雌激活与体细胞克隆胚胎囊胚率（P<0.05）。综上所述，微电极融合法可显著提高绵羊体细胞核移植融合效率，无透明带构建方法不能显著提高克隆胚胎囊胚率，胚胎培养液中添加50 μmol•L^-1维生素C可显著提高绵羊克隆胚胎囊胚率。

关键词: 克隆绵羊, 体细胞核移植, 微电极, 无透明带克隆, 维生素C

Abstract:

The present study was designed to investigate the effects of different nuclear transfer methods and supplementation of culture media with vitamin C on the fusion rate and development of ovine cloned embryos.The abattoir-derived ovaries were collected for materials.After in vitro maturation of oocytes，micro-electrode fusion，zona-free and micro-injection cloning were performed to compare their efficiencies in construction of cloned embryos.Optimal concentration of vitamin C was explored by examining the effects of different concentrations of vitamin C on the parthenogenetic development of oocytes and subsequently applied to in vitro culture of cloned embryos.The results showed that fusion rate obtained from micro-electrode fusion with 25 volts was significantly increased compared with Chamber fusion (P<0.05).Twenty five volts resulted in less dead oocytes than 30 volts in micro-electrode fusion (P<0.05).Zona-free cloning significantly increased the morula rate of cloned embryos (P<0.05)，but failed to increase blastocyst rate significantly (P>0.05).Vitamin C supplementation with the concentration of 50 μmol•L^-1 significantly increased blastocyst rate of ovine parthenogenetic and cloned embryos (P<0.05).Altogether，fusion rate during somatic cell nuclear transfer could be increased by micro-electrode fusion.Blastocyst development of ovine cloned embryos could not be improved by zona-free cloning.Fifty μmol•L^-1 vitamin C was beneficial for in vitro culture of ovine cloned embryos.

Key words: cloned sheep, somatic cell nuclear transfer, micro-electrodes, zona-free cloning, vitamin C

中图分类号:

S826
S814.8

闵江涛，杜卫华，赵学明，郝海生，杭苏琴，朱化彬. 核移植方法和维生素C影响绵羊克隆胚胎体外发育[J]. 畜牧兽医学报, 2014, 45(9): 1410-1416.

MIN Jiang-tao，DU Wei-hua，ZHAO Xue-ming，HAO Hai-sheng，HANG Su-qin，ZHU Hua-bin. Nuclear Transfer Methods and Vitamin C Affect the in vitro Development of Ovine Cloned Embryos[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2014, 45(9): 1410-1416.

参考文献

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