大豆盐胁迫响应NRT基因的鉴定及GmNRT1.5A的克隆和表达分析

Identification of Soybean Salt Stress Responsive NRT Genes and Cloning and Expression Analysis of GmNRT1.5A

硝酸盐转运蛋白(Nitrate Transporters, NRTs)在植物根系NO-3吸收或转运中发挥重要作用,为探究大豆NRTs基因在盐胁迫响应中的功能,本研究从前期盐胁迫处理的转录组数据中鉴定出8个持续响应盐胁迫的差异表达GmNRTs基因。这些GmNRTs基因的启动子上均含有多个与逆境胁迫或植物激素应答相关的元件,推测它们可能与大豆非生物胁迫应答或植物激素调控盐胁迫响应相关。同时,从齐黄34根系中克隆了盐胁迫处理上调倍数最大的基因GmNRT1.5A,并对其进行了生物信息学和表达分析。结果显示:GmNRT1.5A的蛋白质编码区全长为1 794 bp,编码597个氨基酸,蛋白分子质量为66.78 kD,理论等电点为5.85。进化树分析结果表明,GmNRT1.5A与AtNRT1.5同源性最高,含有12个跨膜结构域,含有典型的NRT1s保守结构域,二级结构以α-螺旋和无规则卷曲为主。实时荧光定量PCR分析表明,GmNRT1.5A主要在籽粒和根系中表达,盐胁迫和干旱胁迫显著诱导根系中GmNRT1.5A的表达。此外,GmNRT1.5A的表达受植物激素的调控,ABA和ACC显著诱导其表达,但GA3处理显著抑制GmNRT1.5A表达。本研究表明,GmNRT1.5在响应盐胁迫逆境中发挥重要作用,为探索GmNRTs基因在响应逆境胁迫中的功能提供了新思路,也为深入研究GmNRT1.5基因的功能提供了基础。

Nitrate transporters(NRTs)play an important role in NO-3 uptake or transporters in plant roots.In order to study the function of soybean NRTs genes in response to salt stress,eight differentially expressed GmNRTs genes that respond to salt stress were identified from the transcriptome data of salt stress treatment.The promoters of these GmNRTs genes contain multiple plant stress or plant hormones response elements,suggesting that these genes might be involved in abiotic stress or plant hormone regulation of salt stress response of soybean.At the same time,we cloned GmNRT1.5A from soybean roots,which was the most up-regulated gene under salt stress treatment.The GmNRT1.5A CDs of 1794 bp in length,encoded a 597 amino acid,with molecular weight of 66.78 kD and isoelectric point(pI)of 5.85.Phylogenetic analysis revealed that GmNRT1.5A had a close relationship with AtNRT1.5 and ZmNRT2.5,contained twelve transmembrane domains and typical NRT1s conserved domains,and the secondary structure of GmNRT1.5A was found to be composed ofα-helix and random coils.Real-time quantitative PCR analysis showed that GmNRT1.5A was mainly expressed in seeds and roots.Salt stress and drought stress significantly induced the expression of GmNRT1.5A gene in root.In addition,the expression of GmNRT1.5A was significantly induced by ABA and ACC,while GA3 treatment significantly inhibited the expression of GmNRT1.5A.This study showed that GmNRT1.5A played an important role in response to salt stress,which provides a new idea for exploring the function of NRTs genes in soybean response to stress,and also lays a foundation for in-depth research on the function of GmNRT1.5A gene in soybean.