紫花苜蓿抗逆基因MsDUF的克隆及其功能分析  

Isolation and Functional Analysis of the Stress Resistance Gene MsDUF in Medicago Sativa L.

作　　者：韩博[1,2] 王卫栋[1] 杨培志[1] 张攀[1] 呼天明[1] 

机构地区：[1]西北农林科技大学动物科技学院,陕西杨凌712100 [2]旱区作物逆境生物学国家重点实验室,陕西杨凌712100

出　　处：《中国农业科学》2013年第2期424-432,共9页Scientia Agricultura Sinica

基　　金：国家自然科学基金项目(30901050;31272490);国家"十二五"科技支撑计划(2011BAD17B05)

摘　　要：【目的】克隆紫花苜蓿(Medicago sativa L.)抗逆新基因MsDUF,并对其进行序列特征分析,了解该基因在逆境胁迫下的表达模式。【方法】利用RACE法获得紫花苜蓿MsDUF全长的cDNA序列,对该序列进行生物信息学分析;用基因枪法进行MsDUF的亚细胞定位分析;采用实时荧光定量PCR研究该基因在高含量NaCl和PEG-6000的胁迫下,以及ABA和GA3诱导下的表达模式。【结果】测序结果显示,该基因cDNA全长714 bp,包含一个633 bp的完整开放阅读框,编码210个氨基酸,命名为MsDUF,GenBank登录号为JX183734。氨基酸BLASTP分析表明,MsDUF氨基酸序列与拟南芥、大豆和蒺藜苜蓿等具有53%—98%的相似性,属于DUF4228 superfamily。实时荧光定量PCR研究表明,该基因在逆境胁迫下上调表达。【结论】从紫花苜蓿中克隆了抗逆相关基因MsDUF,发现其定位于细胞质中,实时荧光定量PCR结果分析表明该基因在紫花苜蓿的抗逆反应中起着重要作用。[Objective] This study is aimed to characterize the stress resistance gene MsDUF in Medicago sativa by cloning and analysis of its sequence, and study the function of MSDUF under stress conditions. [Method] The full-length cDNA sequence of MsDUF was isolated by RACE. The obtained cDNA sequence and the deduced amino acid sequence were analysed. Using gene gun in subcellular localization. Real-time quantitative PCR was used to assess the expression of MsDUF in response to NaC1, PEG-6000, ABA and GA3. [Result] The sequencing result showed that the cloned cDNA （designated as MsDUF, GenBank accession No. JX183734） was 714 bp, contains an open reading frame （ORF） of 633bp, and encoding a protein with 210 amino acids. BLASTp search revealed 53% to 98% similarities between the MSDUF protein and those reported MSDUF from other species, such as Arabidopsis thaliana, Glycine max and M.. truncatula, The RT-PCR result showed that MsDUF was upregulatedly in response to stress resistance. [Conclusion] The stress resistance related gene MsDUF was cloned from M. sativa and found that it localized in cytoplasm. RT-PCR analysis showed that the gene MsDUF plays an important role in response to the abiotic stress. It is a novel stress resistance gene in M. sativa.

关 键 词：紫花苜蓿 MsDUF 高渗胁迫 激素诱导 亚细胞定位 功能分析 

分 类 号：S541.9[农业科学—作物学]