‘云香’水仙NtWRKYY1基因的分离与功能分析
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福建省种业创新与产业化工程项目(K8114001B)


Isolation and Function Analysis of NtWRKYY1 Transcription Factor Gene in Narcissus tazetea var. ‘Yunxiang’
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    摘要:

    为明确WRKY转录因子在‘云香’水仙中的功能,该研究以‘云香’水仙为材料,克隆了WRKY基因,命名为NtWRKYY1(GenBank登录号为KX056495)。序列分析显示,NtWRKYY1基因开放阅读框(ORF)长度为510 bp,编码169个氨基酸。多序列比对和系统进化树分析显示,NtWRKYY1编码蛋白含1个WRKY结构域和C2H2锌指结构(Cx4Cx23HxH),与AtWRKY57聚在一起,属于第Ⅱc类WRKY转录因子。组织表达和时空表达分析显示,NtWRKYY1基因在花中表达量高于根和叶,且在花瓣及副冠中的表达量随开花过程(花蕾期、始花期、盛花期、衰败期)呈上升趋势。植物激素和非生物胁迫分析显示,NtWRKYY1基因受脱落酸(ABA)、高温、干旱和盐诱导,受茉莉酸甲酯(JA)抑制,表明NtWRKYY1基因可能在‘云香’水仙花朵的衰老过程中起正调节作用,同时参与‘云香’水仙ABA、JA等激素信号转导及高温、干旱、盐碱等非生物胁迫过程的调控。利用InFusion克隆技术成功构建过表达载体pMDC140NtWRKYY1,并采用农杆菌介导叶盘法转化烟草。RTPCR和GUS染色结果显示,目的基因已成功导入烟草基因组中。

    Abstract:

    WRKY transcription factors play an important adjusting role in the process of plant growth and development, hormone signal transduction and abiotic stress response. To clarify the gene Narcissus tazetea var. ‘Yunxiang’, we cloned the NtWRKYY1 gene (GenBank accession No. KX056495) based on ‘Yunxiang’, analyzed the gene sequence features, evolutionary relationship and expression characteristics, constructed the expression vector before transformed into tobacco. Sequence analysis revealed that the length of NtWRKYY1 gene open reading frame (ORF) is 510 bp, encoding a polypeptide of 169 amino acids. Multiple alignments and phylogenetic analysis showed that NtWRKYY1 protein containing one WRKY consecutive structural domain and C2H2 type zinc finger(Cx4Cx23HxH)belong to Ⅱc subgroup of WRKY transcription factor together with Arabidopsis AtWRKY57. Tissuespecific expression and temporal and spatial expression showed that NtWRKYY1 in ‘Yunxiang’ had a much higher expression in flowers than that in roots and leaves, and a rising express trend in petal and corona of alabastrum stage, early flowering stage, full bloom stage and faded stage. NtWRKYY1 can be induced by abscisic acid (ABA), high temperature, drought and saline, and restrained by methyl jasmonate (JA) through hormones and abiotic stresses analyzing. We can concluded that NtWRKYY1 gene may play a regulating role during the flower senescence process in ‘Yunxiang’, involve in the hormone signal transduction of ABA, JA and the abiotic stress regulation of high temperature, drought and saline at the same time. In addition, we constructed overexpressing vector pMDC140NtWRKYY1 using InFusion cloning technique, transformed into tobacco by the method of Agrobacterium though leaf disc transformation. The carrier of PCR and GUS staining results indicated the resistant plantlets were positive. This study will make a good foundation for further exploring the function of the WRKY transcription factor in ‘Yunxiang’.

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温秀萍,孙申申,杨菲颖,等.‘云香’水仙NtWRKYY1基因的分离与功能分析[J].西北植物学报,2017,37(3):436-444

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  • 在线发布日期: 2017-04-18
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