木薯MeNCED3基因克隆、结构变异及其表达分析

doi:10.13560/j.cnki.biotech.bull.1985.2016.10.018

生物技术通报 ›› 2016, Vol. 32 ›› Issue (10): 148-153.doi: 10.13560/j.cnki.biotech.bull.1985.2016.10.018

木薯MeNCED3基因克隆、结构变异及其表达分析

丁泽红, 付莉莉, 铁韦韦, 颜彦, 胡伟

中国热带农业科学院热带生物技术研究所,海口 571101

收稿日期:2016-04-15 出版日期:2016-10-25 发布日期:2016-10-12
作者简介:丁泽红,男,助理研究员,研究方向：植物分子生物学;E-mail：dingzehong@itbb.org.cn
基金资助:
海南省自然科学基金项目（20163120,20153048）

Cloning and Analysis of Structure Variation and Expression of MeNCED3 Gene in Cassava

DING Ze-hong, FU Li-li, TIE Wei-wei, YAN Yan, HU Wei

Institute of Tropical Bioscience and Biotechnology,Chinese Academy of Tropical Agricultural Science,Haikou 571101

Received:2016-04-15 Published:2016-10-25 Online:2016-10-12

摘要/Abstract

摘要： 旨在揭示木薯MeNCED3基因在干旱等非生物胁迫应答中的作用。用RT-PCR的方法从木薯叶片中克隆MeNCED3基因,用MEGA软件构建进化树;用DnaSP软件分析基因结构变异,用荧光定量PCR技术分析MeNCED3在不同非生物胁迫下的表达特性。结果显示,从木薯中克隆了一个NCED基因MeNCED3,该基因具有1 803 bp的开放阅读框,编码600个氨基酸,含有NCED家族保守结构域。进化树分析表明,MeNCED3与杨树和杞柳中同源基因的亲缘关系较近,序列相似性分别达到83.9%和82.5%。基因结构变异发现,木薯野生种和栽培种之间共有8个错义突变,其中5个可能与MeNCED3的表达量有关。实时荧光定量PCR分析表明,MeNCED3在根中的表达量要远远高于叶片和茎。而且,MeNCED3的表达能被PEG、ABA和NaCl处理显著诱导。因此,MeNCED3在转录水平对木薯渗透胁迫、盐胁迫起调控作用,可作为候选基因进一步研究其在木薯抗旱中的功能。

关键词: 木薯, MeNCED3, 克隆, 结构变异, 表达分析

Abstract: To reveal the roles of MeNCED3 gene in abiotic stresses（e.g.,drought）in cassava. RT-PCR method was applied to clone MeNCED3 gene from cassava leaves,MEGA software was used to construct its phylogenetic tree,DnaSP software was used to analysis its structural variation,and quantitative RT-PCR（qRT-PCR）was used to explore its expression patterns in response to different abiotic stresses. Results showed that a NCED（9-cis-epoxycarotenoid dioxygenase）gene,MeNCED3,was cloned from cassava. This gene had a 1 803 bp open reading frame,encoded 600 amino acids,and contained a NCED conserved domain. Phylogenetic analysis showed that MeNCED3 had close genetic relationship with its homologs from Populus trichocarpa and Salix purpurea,and the sequence similarity was up to 83.9% and 82.5%,respectively. Genetic structural variation revealed that a total of eight mis-sense mutations were identified between wild and cultivated cassava species,of which five may be associated with the different expression levels of MeNCED3. qRT-PCR analysis revealed that MeNCED3 expression was higher in roots than in leaves and stems. In addition,the expression of MeNCED3 was significantly induced by PEG,ABA and NaCl treatments. MeNCED3 played a regulatory role in both salt and osmotic stresses at the transcriptional level and it could be served as a candidate to further study its functions in drought resistance in cassava.

Key words: cassava, MeNCED3, clone, structure variation, expression analysis

丁泽红, 付莉莉, 铁韦韦, 颜彦, 胡伟. 木薯MeNCED3基因克隆、结构变异及其表达分析[J]. 生物技术通报, 2016, 32(10): 148-153.

DING Ze-hong, FU Li-li, TIE Wei-wei, YAN Yan, HU Wei. Cloning and Analysis of Structure Variation and Expression of MeNCED3 Gene in Cassava[J]. Biotechnology Bulletin, 2016, 32(10): 148-153.

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木薯MeNCED3基因克隆、结构变异及其表达分析

Cloning and Analysis of Structure Variation and Expression of MeNCED3 Gene in Cassava

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