斑节对虾泛素结合酶PmUbc基因的克隆及表达分析

Molecular cloning and expression analysis of Ubiquitin-conjugating enzyme gene from black tiger shrimp (Penaeus monodon)

  • 摘要: 以从斑节对虾(Penaeus monodon)肝胰腺转录组数据中筛选的泛素结合酶(Ubiquitin-conjugating enzyme,PmUbc)基因片段为基础,利用SMART-RACE技术获得PmUbc的cDNA全长,并利用生物信息学对其进行了分析。PmUbc基因cDNA全长598 bp,包括编码154个氨基酸的465 bp的开放阅读框(ORF)、17 bp的5′非编码区(UTR)和116 bp的3′UTR。Blast分析显示,PmUbc与凡纳滨对虾(Litopenaeus vannamei)等物种的Ubc具有较高的同源性。利用实时荧光定量PCR技术研究了PmUbc mRNA在不同组织和卵巢发育各时期的相对表达量和变化模式。结果表明,PmUbc mRNA在斑节对虾的脑、胸神经、心脏、肝胰腺、卵巢、肌肉、血淋巴中均有表达;PmUbc基因在卵巢发育Ⅲ期表达量最高,其次是Ⅰ和Ⅳ期。结果显示,PmUbc基因在斑节对虾卵巢发育过程中可能具有重要的作用,该实验结果为斑节对虾卵巢发育调控的分子机制研究提供一定的理论依据。

     

    Abstract: We cloned the complete cDNA sequence of Ubiquitin-conjugating enzyme gene from black tiger shrimps (Penaeus monodon) (PmUbc) by rapid amplification of cDNA ends (RACE) methods based on the partial sequence of PmUbc obtained from the hepatopancreas transcriptome database. The cDNA of PmUbc was 598 bp, including 154 amino acids with an open reading frame (ORF) of 465 bp, 17 bp of 5′ untranslation region (UTR) and 116 bp of 3′ UTR. Blast analysis shows that PmUbc shared high identity with Ubiquitin-conjugating enzyme of Litopenaeus vannamei. Real time quantitative PCR analysis reveals that PmUbc was constitutively expressed in seven selected tissues (brain, thoracic nerve, heart, hepatopancreas, ovary, muscle, hemolymph). The PmUbc mRNA expression reached the maximum at Stage Ⅲ of ovarian development, followed by StageⅠand Ⅳ. It is concluded that PmUbc might play an important role in ovarian development of P.monodon.

     

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