• 中文核心期刊
  • CSCD来源期刊
  • 中国科技核心期刊
  • CA、CABI、ZR收录期刊

木薯MeERF1基因克隆及其表达分析

Cloning and Expression of Cassava MeERF1

  • 摘要:
    目的 乙烯响应因子(ethylene responsive factor,ERF)在植物生长发育、非生物胁迫以及抗氧化系统中均具有重要功能。克隆木薯MeERF1基因并进行亚细胞定位和表达分析,为深入研究其潜在功能提供参考。
    方法 利用PCR扩增从木薯SC8品种中获得特异性MeERF1基因CDS序列(全长1425 bp),综合运用生物信息学方法对序列进行功能注释与结构预测,并通过农杆菌介导的瞬时转化体系,检测MeERF1蛋白的亚细胞定位情况。同时检测木薯不同组织以及脱落酸(abscisic acid,ABA)处理块根后不同时间点MeERF1基因的相对表达量。
    结果 PCR克隆获得的MeERF1基因CDS序列与Phytozome数据库收录的参考核苷酸序列(登录号:Manes.13G148300)一致性高达100%,功能预测显示该CDS编码的蛋白由474个氨基酸组成(分子量52.7 kDa,理论pI 6.28),属于酸性蛋白类型。预测分析表明该蛋白含有90.30%的无规则卷曲结构、6.96%的α-螺旋构象和2.74%的延伸链片段,亚细胞定位试验证实了MeERF1蛋白定位于细胞核。通过多序列比对及系统进化分析发现,MeERF1蛋白包含一个关键的AP2/ERF家族保守结构域,该结构域特异性结合GCC/box,证明其作为转录因子发挥生物学功能。此外,该蛋白与巴西橡胶树ERF蛋白的氨基酸序列相似度最高(80.33%)。组织表达模式分析表明MeERF1基因在木薯的不同组织中均有表达,其中在须根中表达最为显著,远高于其他组织类型,茎部次之。MeERF1基因在采后不同时间的块根切片中受诱导表达,且相对表达在ABA处理12 h后显著提高,达到峰值。
    结论 MeERF1基因属于AP2/ERF类转录因子,定位于细胞核,主要在须根中表达,在块根采后生理衰变过程中受诱导,ABA处理显著增强诱导作用,表明MeERF1可能通过响应ABA信号在木薯块根采后生理衰变过程中发挥功能。

     

    Abstract:
    Objective The ethylene responsive factor (ERF) gene in cassava, MeERF1, was cloned for functional analysis.
    Method ERF crucially related to the growth, development, abiotic stress response, and antioxidant regulation of cassava SC8 cultivar was cloned. CDS sequence of MeERF1 gene was obtained by PCR, functional annotation and structural prediction performed by bioinformatics methods, subcellular localization detected by Agrobacterium-mediated transient transformation, and relative expressions in tissues at different sampling times after abscisic acid (ABA) treatment of tubers determined.
    Result The 1,425bp long CDS sequence of MeERF1 as determined was perfectly consistent with what is shown for the reference in the Phytozome database (accession number: Manes.13G148300). An acidic protein, it was composed of 474 amino acids with a molecular weight of 52.7kDa and a pI of 6.28. Structurally, the protein contained 90.30% of irregular coils, 6.96% of α-helixes, and 2.74% of extended strands. It mainly aggregated in the nuclear region, indicating a high possibility of being a transcription factor. In addition, the multiple sequence alignment and phylogenetic analysis showed its inclusion of a key AP2/ERF conserved family domain specifically bound to GCC/box like a transcription factor. It also shared the highest similarity at 80.33% on amino acid sequence with the ERF protein of Brazilian rubber tree. In various tissues of a cassava plant, the expression of MeERF1 was significantly higher in the hairy fine roots, followed by the stems, than in other organs. And the expression peaked after 12h of ABA induction.
    Conclusion The cloned MeERF1 belonged to the AP2/ERF transcription factor that was tissue-specific and responded to ABA. It was plausible that it involved in the regulation of post-harvest physiological deterioration of cassava roots.

     

/

返回文章
返回