珍稀泌盐植物长叶红砂两个WRKY转录因子的克隆及表达分析

doi:10.11686/cyxb20140415

摘要/Abstract

摘要： 根据长叶红砂转录组数据信息,选取在盐胁迫处理后表达上调明显且注释结果为WRKY转录因子的两个cDNA片段。通过cDNA末端快速扩增技术(rapid amplification of cDNA ends,RACE),从长叶红砂中克隆获得2个WRKY基因。在NCBI数据库比对与拟南芥AtWRKY33同源性分别为79%和87%,因此命名为RtWRKY33-1和RtWRKY33-2。其中RtWRKY33-1的cDNA全长2163 bp,开放阅读框(open reading frame,ORF)长度为1681 bp,编码573个氨基酸;RtWRKY33-2的cDNA全长2155 bp,开放阅读框长度为1776 bp,编码591个氨基酸。氨基酸序列分析表明这两个基因均具有两个WRKY结构域,属于典型的I类WRKY转录因子。蛋白结构预测分析发现,两个蛋白的一级结构和二级结构在WRKY结构域的氨基酸序列和结构特性的相似性较高,但是在非保守结构域部分,尤其是N末端(1~80 aa之间)、第1个WRKY结构域之前(190~240 aa之间)和两个WRKY结构域之间(430~450 aa之间)等位置的差异明显,可能对两个基因功能有一定影响。半定量RT-PCR分析表明,4种非生物胁迫均能诱导这两个基因的表达,但是盐、冷和ABA对RtWRKY33-2诱导尤为明显,同时RtWRKY33-2对干旱胁迫的响应更快,因此这两个基因在长叶红砂抵御非生物胁迫的应答反应中发挥的作用可能不同。本研究为深入探索WRKY33转录因子在长叶红砂中的作用机制奠定了基础。

Abstract: Reaumuria trigyna is an endangered small shrub thriving in vast arid areas of Inner Mongolia. This species has developed a distinct morphology and physiology to adapt to the semi-desert environment. The WRKY gene family are plant-specific transcription factors which play important roles in the regulation of plant development,metabolism,and diverse biotic and abiotic stresses. Based on R. trigyna transcriptome data,two cDNA fragments (annotated as WRKY transcription factors) which were significantly upregulated after salt treatment were selected. Using rapid amplification of cDNA ends (RACE),these two WRKY transcription factors were cloned from R. trigyna. Blasted in the NCBI database,the homology to Arabidopsis AtWRKY33 were 79% and 87%,so they were named RtWRKY33-1 and RtWRKY33-2 (GenBank accession number KF421158 and KF421159),respectively. The full-length of the RtWRKY33-1 was 2163 bp and included a 1681 bp open reading frame (ORF) which encoded 573 amino acids. The full-length of RtWRKY33-2 was 2155 bp and included a 1776 bp ORF encoding 591 amino acids. Amino acid sequence analysis indicated that they were typical of class I WRKY transcription factors. Protein structure prediction analysis found that the primary structure and secondary structure in the WRKY domain of the amino acid sequence had structures characteristic of high similarity. However,in the non-conserved domains,especially the N-terminus (1 to 80 aa range),before the first WRKY domain (190 to 240 aa range) and between two WRKY domains (430 to 450 aa range),there were significant differences that may influence the function of these two genes. Semi-quantitative RT-PCR analysis showed that four abiotic stresses can induce the expression of these two genes. However,RtWRKY33-2 was mainly inducted by salt,cold and ABA,while RtWRKY33-2 responded faster to drought stress. These two genes in R. trigyna may play a different rolein abiotic stress responses. This research is a basis for study on the regulation mechanism of WRKY33 transcription factors in R. trigyna.

中图分类号:

Q943.2

王佳，郑琳琳，顾天培，王学峰，王迎春. 珍稀泌盐植物长叶红砂两个WRKY转录因子的克隆及表达分析[J]. 草业学报, 2014, 23(4): 122-129.

WANG Jia,ZHENG Lin-lin,GU Tian-pei,WANG Xue-feng,WANG Ying-chun. Cloning and expression analysis of two WRKY transcription factors from the rare recretohalophyte Reaumuria trigyna[J]. Acta Prataculturae Sinica, 2014, 23(4): 122-129.

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