论文部分内容阅读
为探讨太子参环肽成分与品质形成的分子机制,利用生物信息学方法从太子参转录组数据库中筛选出太子参类胡萝卜素双加氧裂解酶(carotenoid cleavage dioxygenases,CCDs)新基因,并设计引物对其进行全长扩增及克隆、生物信息学分析和基因表达分析。克隆得到的4条CCDs序列全长分别为1 617,1 461,1 746,1 875 bp,并根据其功能依次命名为Ph CCD1,Ph NCED2,Ph NCED3,Ph CCD4。序列分析结果表明,4个基因均含有REP65结构域,且均含有与亚铁离子结合的位点。系统进化分析表明,Ph NCED2,Ph NCED3聚为NCEDs一支,Ph CCD1,Ph CCD4聚为CCDs一支,且与拟南芥Arabidopsis thaliana的At CCDs家族相比,Ph CCD1与At CDD1同源性最高,Ph CCD4与At CCD4同源性最高,Ph NCED2,Ph NCED3与At NCED3同源性最高。实时荧光定量分析显示Ph CCD1和Ph CCD4 2个基因的表达主要在地上部分,其中Ph CCD1在叶中的表达量最高,Ph CCD4在茎叶具有高表达,可能参与太子参类胡萝卜素的生物合成;Ph NCED2和Ph NCED3 2个基因的表达主要在地下部分,其中Ph NCED2在须根中表达量最高,Ph NCED3在块根木部和皮部表达量较高,可能是脱落酸生物合成的关键酶基因。研究首次得到太子参CCDs基因,为进一步阐明太子参植株对于外界胁迫的应答机制,进而探讨太子参品质形成的生物途径奠定基础。
In order to explore the molecular mechanism of the composition and quality of rhizoma polygonatum, bioinformatics methods were used to screen novel genes of heterotrophic heterophylla carotenoid cleavage dioxygenases (CCDs) Primer for its full-length amplification and cloning, bioinformatics analysis and gene expression analysis. The four cloned CCDs sequences were 1617, 1461, 1746 and 1875 bp, respectively, and named Ph CCD1, Ph NCED2, Ph NCED3 and Ph CCD4 according to their functions. The results of sequence analysis showed that all four genes contained REP65 domain, and all contained the sites that bound to ferrous ion. Phylogenetic analysis showed that Ph NCED2 and Ph NCED3 clustered into one NCEDs, Ph CCD1 and Ph CCD4 clustered into one CCDs, and Ph CCD1 had the highest homology with At CDD1 compared with At CCDs family of Arabidopsis thaliana Ph CCD4 had the highest homology with At CCD4 and Ph NCED2 and Ph NCED3 had the highest homology with At NCED3. Real-time fluorescence quantitative analysis showed that the expression of Ph CCD1 and Ph CCD4 genes were mainly in the aerial part, of which Ph CCD1 was highest in leaves and Ph CCD4 was highly expressed in stems and leaves, which may be involved in the biosynthesis of heterophylla carotenoids ; Ph NCED2 and Ph NCED3 mainly expressed in underground parts, of which Ph NCED2 was highest in the fibrous roots and Ph NCED3 was higher in the roots and husks, which may be the key enzyme of abscisic acid biosynthesis . This study firstly obtained the CCDs gene of heterophylla heterophylla, which lays the foundation for further elucidation of the response mechanism of heterophylla heterophylla to external stress, and then to explore the biological pathway of heterophylla heterophylla formation.