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超表达Cdc20基因不影响牛卵母细胞第一极体排出 被引量：1

Over-expression of Cdc20 Gene Has No Effect on Bovine Oocytes First Polar Body Extrusion
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摘要 CDC20(cell division cycle 20)是后期促进复合物(anaphase-promoting complex,APC)的共激活剂之一,也是纺锤体组装检查点(spindle assembly checkpoint,SAC)的靶点,在细胞周期调控中扮演重要角色.为探讨Cdc20在第一极体排出(first polar body extrusion,PBE I)中的作用,Cdc20基因被成功克隆并构建了真核表达载体pCdc20-Venus,随后用T7 Mmessage Mmachine Kit(Ambion)以线性化pCdc20-Venus为模板体外转录(in vitro transcription)获得带帽的Cdc20-Venus mRNA,将Ccdc20-Venus mRNA显微注射到体外培养的牛卵母细胞胞质中进行超量表达.结果表明,真核表达载体pCdc20-Venus转染HeLa细胞后能够正常表达,绿色荧光在细胞核周围呈弥散状分布;将Cdc20-Venus mRNA注射到牛卵母细胞胞质后,胞质内有绿色荧光出现.Cdc20-Venus mRNA注射组卵母细胞的PBE I率(48.9%)与Venus mRNA注射组卵母细胞的PBE I率(50.9%)和未注射组卵母细胞的PBE I率相比均没有显著差异(P>0.05).通过Cdc20在牛卵母细胞内的超量表达,结果显示,超量表达Cdc20基因对牛卵母细胞PBE I没有显著影响(P>0.05). As one of the co-activator of anaphase-promoting complex （ APC）, cell division cycle 20 （CDC20） protein also functions as the target of the spindle assembly checkpoint （SAC）, which is essential for the cell cycle regulation. To investigate the function of Cdc20 during the first polar body extrusion （PBE I）, Cdc20 CDS was cloned and eukaryotic expression vector pCdc20-Venus was constructed. Using the linear pCdc20-Venus as template, the capped Cdc20-Venus mRNA was synthesized via T7 Mmessage Mmachine Kit （Ambion）. Cdc20 over-expression was performed by microinjection of Cdc20-Venus mRNA into the cytoplasm of bovine oocytes. The results showed that Venustagged Cdc20 dispersed around the nucleus in HeLa cells. In bovine oocytes, the fluorescence appeared in the whole cytoplasm. However, the PBE I rate in over-expressed group （48.9%） is not significant, compared to Venus mRNA injection group （50.9%） and non-injection group （51.1%）. Our study demonstrated that the over-expression of Cdc20 in bovine oocytes does not affect the PBE I rate （P 〉 0. 05）.
作者 杨文琳 安鹏 李伟 赵贵民 史芸安 雷安民
机构地区 西北农林科技大学动物医学院 陕西省干细胞工程技术研究中心 山东省农业科学院奶牛研究中心
出处 《中国生物化学与分子生物学报》 CAS CSCD 北大核心 2012年第6期546-554,共9页 Chinese Journal of Biochemistry and Molecular Biology
基金 国家自然科学基金项目(No.31172280)~~
关键词 牛 Cdc20 卵母细胞 超表达 减数分裂 第一极体排出率 bovine Cdc20 oocytes over-expression meiosis PBE I rate
分类号 Q78 [生物学—分子生物学]
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参考文献26

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二级参考文献17

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共引文献6

同被引文献16

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中国生物化学与分子生物学报
2012年 第6期

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