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草业学报  2014 

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, PP. 122-129

Keywords: 长叶红砂,WRKY转录因子,克隆,表达分析

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Abstract:

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(rapidamplificationofcDNAends,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2163bp,开放阅读框(openreadingframe,ORF)长度为1681bp,编码573个氨基酸;RtWRKY33-2的cDNA全长2155bp,开放阅读框长度为1776bp,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(1~80aa之间)、第1个WRKY结构域之前(190~240aa之间)和两个WRKY结构域之间(430~450aa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。

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