甘蔗钾转运蛋白基因SsHAK2的克隆及表达特性分析  

Cloning and Expression Analysis of Potassium Transporter SsHAK2 in Sugarcane(Saccharum species hybrid)

作　　者：凌秋平[1] 曾巧英[1] 胡斐 吴嘉云[1] 樊丽娜[1] 李奇伟[1] 齐永文[1] 

机构地区：[1]广东省生物工程研究所(广州甘蔗糖业研究所)广东省甘蔗改良与生物炼制重点实验室,广州510316

出　　处：《农业生物技术学报》2017年第3期378-385,共8页Journal of Agricultural Biotechnology

基　　金：国家甘蔗产业技术体系(CARS-20-1-4);广东省科技计划项目(No.2014B070705002;No.2014A040401033;No.2014B090907006);广东省科学院科研平台环境与能力建设专项资金项目(No.2016GDASPT-0306);亚热带农业生物资源保护与利用国家重点实验室开放课题

摘　　要：甘蔗(Saccharum species hybrid)是最重要的糖料作物,由于其生育周期长,生物量大,对钾的需求量也大。南方是我国甘蔗的主要种植区域,由于土壤本身特性,有效钾含量低,限制了甘蔗的生产。因此,培育耐低钾甘蔗品种是提高甘蔗钾吸收效率的有效途径之一。本研究以甘蔗品种新台糖22号为材料进行低钾胁迫处理,利用RT-PCR技术从其根系中克隆得到钾转运蛋白基因,命名为SsHAK2(Gen Bank登录号:KM98738)。该基因全长为2 798 bp,包含一个完整的2 352 bp的ORF,编码784个氨基酸,相对分子量为87.602 k D,等电点为8.85,预测其为碱性蛋白。SsHAK2苷酸序列包含了12个跨膜结构域(S1~S12),有80%的概率定位在细胞质膜上。同时,该基因还包含3个保守结构域,分别为钾转运蛋白结构域和氨基酸转运蛋白结构域。SsHAK2基因与玉米(Zea mays)、大麦(Hordeum vulgare)、水稻(Oryza sativa)等其他作物中HAK基因具有高度的同源性,一致的核苷酸比例在52%~95%。q PCR分析结果表明,在低钾、干旱和盐胁迫下,SsHAK2的表达都发生改变。在低钾胁迫条件下处理96 h,该基因相对表达量最大,约达到对照的1.70倍;在盐胁迫条件下处理48 h,该基因的相对表达量出现显著上调,96 h相对表达量最高,约为对照的4.37倍。在干旱胁迫12 h,该基因的相对表达量达到最高,约为对照的4.07倍。干旱胁迫处理24 h,该基因表达量约为对照的1.69倍。但是,在干旱胁迫24到48 h过程中表达由下调转为上调。干旱胁迫48 h后,该基因的表达迅速下调,96 h时该基因表达量最低,约为对照表达量的1/4。q PCR分析结果表明该基因在低钾、干旱和盐胁迫下发挥重要的调控作用。本研究结果为进一步研究甘蔗钾吸收分子机制提供基础。Sugarcane （Saccharum species hybrid） is the most important crop for the production of sugar. Due to its high biomass production and lengthy growing season, sugarcane needs to absorb a large amount of potassium （K） throughout its life cycle. In the southern China where sugarcane is cultivated, most of the soil have low content of available K, which limits the production of sugarcane. To improve the productivity, one of the efficient means is to improve sugarcane cultivars that have efficient uptake capacity of K and hence are tolerant to low K. This study was conducted in which a major commercial cultivar, ROC22, was subject to low K stress. From the experiment, a potassium transporter gene, named SsHAK2 （GenBank accession number： KM98738）, was cloned by reverse transcription-polymerase chainreaction （RT-PCR） from total RNA of sugarcane roots. The full length of the SsHAK2 gene was 2 798 bp, and contained a complete open reading frame of 2 352 bp encoding a protein with 784 amino acids. The molecular weight of SsHAK2 was 87.602 kD and the isoelectric point of 8.85, as the alkaline protein. It contained twelve transmembrane segmentand（S1~S12） and the possibility of subcellular localization in plasma membrane was 80%. The deduced polypeptide of SsHAK2 had three transmembrane domains, including potassium transporter domain and amino acid transporter domain. The homology analysis indicated that SsHAK2 shared sequence homology with other members of HAK family in plant, such as Zea mays, Oryza sativa, Hordeum vulgare, with the level of sequence identity ranging from 52% to 95%. The changes in expression of SsHAK2 under low K, drought and salt stresses were detected by qPCR analysis. Under low K stress condition, the gene had the highest inducible expression level at 96 hours, which was 1.70-fold than that of control. Under salt stress, the expression level began to increase rapidly after 48 hours stress of salt and also reached the highest at 96 hours or 4.37-fold than that of control. Under drough

关 键 词：甘蔗 钾转运蛋白 SsHAK2 基因克隆 基因表达 

分 类 号：S566.1[农业科学—作物学]