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核农学报  2014 

绵羊卵巢microRNA的克隆及表达分析

DOI: 10.11869/j.issn.100-8551.2014.12.2184, PP. 2184-2191

Keywords: 绵羊,卵巢,microRNA,克隆,生物信息学,实时荧光定量PCR

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Abstract:

miRNA(microRNA)是广泛存在于真核生物中长约18~25个核苷酸的一类非编码RNA,调控基因转录后表达。目前,对miRNA的鉴定、功能分析及机理调控的研究已成为生物学领域的热点。为丰富绵羊miRNA数据库、研究miRNA对季节性繁殖动物繁殖活动的调控作用及调控机理提供基础。本研究通过克隆法构建绵羊卵巢miRNAcDNA文库,并进行生物学分析;利用RealtimePCR检测miRNA在卵巢组织中的表达。试验成功构建绵羊卵巢miRNA的cDNA文库,获得30条绵羊miRNA序列。miRBase数据库序列比对,20条miRNA序列与绵羊已知miRNA同源保守,鉴定为绵羊已知miRNA。其中oar-miR-3955-5p在其它物种中未注释,鉴定为绵羊特异性miRNA。通过比对及二级结构分析鉴定了绵羊10条新miRNA,其中ovis-aries-ovary-m0001-5p只与牛bta-miR-2285具有一定的同源性。RealTimePCR结果表明10个miRNA(5个已知miRNA及5个新miRNA)在绵羊卵巢组织中均有表达,但ovis-aries-ovary-m0003-3p的表达量显著高于其它9个,推测该新miRNA对绵羊卵巢发育或功能发挥具有重要的调控作用。试验结果对绵羊miRNA的进一步研究具有一定的理论指导和现实意义。

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