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木薯NAC转录因子Rd26基因克隆及表达
作者: 丁泽红   付莉莉   铁韦韦  伟* 
单位: (中国热带农业科学院 热带生物技术研究所 海口 571101) 
关键词: 木薯 NAC转录因子 Rd26基因 克隆 表达 
分类号:S533
出版年,卷(期):页码:2016,47(11):1822-1826
摘要:

【目的】克隆木薯NAC转录因子Rd26基因(MeRd26)并检测其在干旱胁迫下的表达量,为Rd26基因抗旱调控机制研究打下基础。【方法】利用反RT-PCR克隆木薯叶片中的MeRd26基因,对其进行序列比对及系统发育进化树构建,研究其在栽培种Ku50和野生种W14间的变异情况,并用实时荧光定量PCR(qPCR)检测PEG-6000干旱胁迫下的MeRd26基因表达量。【结果】从木薯叶片中克隆获得的MeRd26基因,长度为1288 bp,包含1041 bp的开放阅读框,编码346个氨基酸,且含NAC保守结构域。系统发育进化树分析结果表明,MeRd26蛋白与杨树(Potri.011G123300.1)和杞柳(SapurV1A.0127s0020.1)的Rd26蛋白亲缘关系较近。基因结构变异分析结果显示,MeRd26基因有33个SNP位点和7个InDel位点,且大部分变异分布在非编码区及最后一个外显子的后半区域。基因表达检测结果显示,在正常大田种植条件下,栽培种Ku50叶片的MeRd26基因表达量是野生种W14的130倍,但在根中表达量差异较小;在干旱胁迫下,栽培种Ku50未展开叶、老叶和根中MeRd26基因表达被快速诱导,表达量随胁迫时间的延长而增加,在根中表达量最高,但在第1片完全展开叶中,MeRd26基因的表达被抑制,其表达量明显降低。【结论】MeRd26基因在转录水平上参与了木薯抗干旱胁迫反应,可作为候选基因用于木薯抗旱机制研究。

【Objective】The present experiment was conducted to clone NAC transcription factor Rd26 gene(MeRd26) from Manihot esculenta Crantz and detect its expression level under drought stress,in order to provide a foundation for further research on regulation mechanism of MeRd26 gene in response to drought. 【Method】MeRd26 gene was cloned from M. esculenta leaf by RT-PCR,and its sequence was aligned with sequences of other species,then their phylogenetic tree was constructed. Subsequently,its structural variation between wild variety W14 and cultivated variety Ku50 was revealed,and its expression level under PEG-6000 drought stress was detected by quantitive real-time PCR(qPCR). 【Result】MeRd26 gene was cloned successfully from M. esculenta leaf,with length of 1288 bp,containing open reading frame(1041 bp),encoding 346 amino acids,containing NAC conserved domain. Phylogenetic tree showed that,MeRd26 protein had close genetic relationship with Rd26 protein of Populus trichocarpa(Potri.011G123300.1) and Salix purpurea(SapurV1A.0127s0020.1). Gene structure variation analysis revealed that MeRd26 gene had 33 SNPs and 7 InDels,and most of these variations were located in the non-coding region and second half of last exon. Expression analysis showed that,MeRd26 gene in leaf of cultivated variety Ku50 was expressed 130 times as that in wild variety W14,while difference was small in root. Under drought stress,expression of MeRd26 gene was rapidly increased in folded leaf,bottom leaf and root with increase of drought stress time,especially in root with the highest expression level. On the contrary,expression of MeRd26 gene was inhibited and reduced obviously in the first fully expanded leaf. 【Conclusion】MeRd26 gene is involved in drought-resistant reaction at transcriptional level,so it can be served as a candidate gene to further study its role in drought-resistant mechanism of M. esculenta.

基金项目:
海南省自然科学基金项目(20163120,314123)
作者简介:
*为通讯作者,胡伟(1982-),博士,主要从事植物分子生物学研究工作,E-mail:huwei2010916@126.com。丁泽红(1982-),主要从事植物分子生物学研究工作,E-mail:dingzehong@itbb.org.cn
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