筇竹钾离子通道QtSKOR1基因的克隆及表达分析

Cloning and expression analysis of QtSKOR1 gene of Qiongzhuea tumidinoda potassium ion channel

【目的】外向整流型钾离子通道SKOR(Stelar K+outward rectifier)家族是参与植物钾离子转运和分配的重要通道,且K+在植物生理过程和响应非生物胁迫中有决定性作用。旨在研究外向整流型钾离子通道SKOR在富含钾元素的筇竹(Qiongzhuea tumidinoda)发育及受到盐胁迫过程中的功能。【方法】利用RACE技术从筇竹的幼苗中获得SKOR基因并命名为QtSKOR1(基因号:MT078984),并对其进行生物信息分析及表达特征分析。【结果】QtSKOR1基因开放阅读框(1923 bp)编码了641个氨基酸。QtSKOR1蛋白存在环核苷酸(cNMP)和锚蛋白重复结构域(ANK)属于Shake亚家族,其相对分子质量为157.27 kD,理论等电点为4.94,含有3个跨膜区但不存在信号肽,属于疏水性蛋白。亚细胞定位分析表明QtSKOR1蛋白主要定位于线粒体(30.4%)和细胞质(26.1%)中。同源分析和进化分析显示,QtSKOR1与二穗短柄草(Brachypodium distachyum)和玉米(Zea mays)的同源性较高,分别为89.06%和87.91%。qPCR结果显示,QtSKOR1基因在筇竹所有组织中均有表达,且表达量从高到低依次为叶、根、茎、笋。与对照相比,随着钾胁迫时间的增加,QtSKOR1的表达量在根中显著增加,而在茎叶中显著降低。随着钠胁迫时间的增加,QtSKOR1的表达量在根茎叶中均呈下降趋势。【结论】QtSKOR1基因属于Shake亚家族,均参与了筇竹各个组织特别是叶的钾运输。同时,在钾胁迫下QtSKOR1基因在根中发挥了积极作用。

[Objective]The SKOR(Stelar K+outward rectifier)family of potassium channels is an important channel involved in the transport and distribution of potassium ions.Furthermore,K+plays a decisive role in plant physiological process and response to abiotic stress.The aim is to investigate the function of outward rectifying potassium ion channel SKOR in the development and salt stress process of potassium rich Q.tumidinoda.[Method]SKOR gene was cloned from the seedlings of Q.tumidinoda by RACE technique and called QtSKOR1(gene number:MT078984).Then analyzed its biological information and gene expression characteristics.[Result]The results showed that the open reading frame(1923 bp)of the QtSKOR1 gene encodes 641 amino acids.QtSKOR1 protein has cNMP and ankyloprotein repeat domain(ANK),belonging to Shake subfamily,and its relative molecular weight is 157.27 kD.The theoretical isoelectric point is 4.94,it contains 3 transmembrane regions,but there is no signal peptide,and it is a hydrophobic protein.In addition,Subcellular localization analysis showed that QtSKOR1 protein mainly existed in mitochondria(30.4%)and cytoplasm(26.1%).Homology and evolutionary analyses showed that QtSKOR1 had high homology with Brachypodium distachyum and Zea mays,with 89.06%and 87.91%,respectively.The qPCR results showed that the QtSKOR1 gene was expressed in all tissues of Q.tumidinoda,and the expression levels was in the order of leaf>root>stem>shoot from high to low.Compared with the control,with the increase of potassium stress time,the expression level of QtSKOR1 was significantly increased in roots and significantly decreased in the stems and leaves.With the increase of sodium stress time,the expression level of QtSKOR1 showed a decreasing trend in roots,stems,and leaves.[Conclusion]The QtSKOR1 gene belongs to the Shake subfamily and is involved in potassium transport in various tissues of Q.tumidinoda,especially in leaves.At the same time,the QtSKOR1 gene played an active role in root under potassium stress.