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郝国礼,王淑梅,刘雅贤,王海洋,于海泉.2015.多能性转录因子OCT4和SOX2在体外发育的小鼠2-细胞胚胎的表达与定位.动物学杂志,50(2):176-184.
多能性转录因子OCT4和SOX2在体外发育的小鼠2-细胞胚胎的表达与定位
The Expression and Localization of Pluripotency Transcription Factors OCT4 and SOX2 in 2-Cell Mouse Embryos Developed In vitro
投稿时间:2014-07-03  修订日期:2015-03-11
DOI:DOI: 10.13859/j.cjz.201502003
中文关键词:  小鼠胚胎  合子基因组激活  2-细胞阻滞  Oct4  Sox2
英文关键词:Mouse embryo  Zygotic genome activation (ZGA)  2-cell block  Octamer-binding transcription factor 4 (OCT4)  Sex determining region Y box protein 2 (SOX2)
基金项目:内蒙古自然科学基金重大项目(No. 2012ZD04),国家自然科学基金项目(No. 30860185 );* 通讯作者,E-mail: haiquan_yu@163.com;第一作者介绍 郝国礼,男,硕士研究生;研究方向:生殖生物学;E-mail: haoguoli123@163.com。 内蒙古大学 哺乳动物生殖生物学及生物技术教育部重点实验室 呼和浩特 010070
作者单位E-mail
郝国礼 内蒙古大学 哺乳动物生殖生物学及生物技术教育部重点实验室 呼和浩特 010070 haoguoli123@163.com 
王淑梅 内蒙古大学 哺乳动物生殖生物学及生物技术教育部重点实验室 呼和浩特 010070  
刘雅贤 内蒙古大学 哺乳动物生殖生物学及生物技术教育部重点实验室  
王海洋 内蒙古大学哺乳动物生殖生物学及生物技术教育部重点实验室  
于海泉 内蒙古大学哺乳动物生殖生物学及生物技术教育部重点实验室 haiquan_yu@163.com 
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中文摘要:
      研究的科学问题:探讨多能性转录因子OCT4和SOX2在昆明小鼠(Mus musculus)2-细胞胚胎发育过程中与2-细胞胚胎阻滞发生的相关性。研究方法:本研究应用实时荧光定量PCR 技术检测了小鼠卵母细胞及在M16培养液中培养的不同发育阶段体外受精胚Oct4和Sox2基因的表达,并利用实时荧光定量PCR 和免疫荧光技术比较了2-细胞期胚、2-细胞阻滞胚和4-细胞期胚的OCT4和SOX2的表达与定位。采用ANOVA对实验所得的数据进行分析,P<0.05被认为是具有显著性差异。研究结果:2-细胞期胚只有24.8%发育成4-细胞期胚,75.2%的2-细胞胚发生了阻滞。Sox2和Oct4的mRNA在MII期卵母细胞、原核胚、2-细胞、4-细胞、桑椹胚和囊胚中都有表达。Oct4 mRNA 的表达水平在4-细胞期胚显著高于2-细胞期胚和2-细胞阻滞胚(P<0.05),Sox2 mRNA 的表达水平在2-细胞期胚显著高于2-细胞阻滞胚和4-细胞期胚(P<0.05),而后两者之间没有差异(P< 0.05)。OCT4蛋白在2-细胞期胚和4-细胞胚中与核共定位,但在2-细胞阻滞胚中弥散存在于胞质中。SOX2蛋白在以上3类胚胎中始终定位于细胞核。研究结论:转录因子OCT4和SOX2的表达与定位与小鼠2-细胞胚胎发育阻滞相关,母源性SOX2表达的维持对胚胎合子基因组激活(ZGA)的发生具有重要作用,母源性OCT4的异常定位可能影响了合子基因组激活相关基因的激活,而合子中Oct4的表达影响合子基因组激活后胚胎的发育。
英文摘要:
      The key scientific problem: the pluripotency transcription factors OCT4 and SOX2 are related to 2-cell block, which usually occurs in mouse ( Mus musculus) embryos cultured in vitro. The main research methods: the expression of Oct4 and Sox 2 was detected by real-time quantitative PCR in oocytes and embryos which were cultured in M16 medium. Meanwhile, the expression and localization of OCT4 and SOX2 were compared among 2-cell stage embryos, 2- cell arrested embryos and 4-cell stage embryos by real-time quantitative PCR and immunofluorescence analysis. Statistical analyses of real-time quantitative PCR results were conducted using an analysis of variance (ANOVA), and a difference of P<0.05 was considered significant. The research results: 24.8% of 2-cell embryos reached the 4-cell stage, and 75.2% of 2-cell embryos were blocked (Fig. 1). Oct4 and Sox2 were expressed throughout MII oocytes, pronuclear zygotes, 2-cell embryos, 4-cell embryos, morulae and blastocysts (Fig. 2a, b). Oct4 was expressed higher in 4-cell embryos than in 2-cell embryos and 2-cell arrested embryos (Fig. 2c) ( P< 0.05). Sox2 was expressed higher in 2-cell embryos than in 4-cell embryos and 2-cell arrested embryos (P < 0.05), but there was no significant difference between 4-cell embryos and 2-cell arrested embryos (Fig. 2d) (P < 0.05). OCT4 was co-localized with chromatin in 2-cell and 4-cell nuclei, while diffused in cytoplasm instead of in nuclei in 2-cell arrested embryos (Fig. 3a). SOX2 was co-localized with chromatin in all groups (Fig. 3b). The research conclusion: the expression and localization of OCT4 and SOX2 is related to 2-cell block in mouse embryos. The stable expression of maternal SOX2 plays an important role during zygotic genome activation (ZGA). The abnormal localization of maternal OCT4 may affect the activation of ZGA-related genes; moreover, the expression of zygotic Oct4 may impact embryo development after ZGA.
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