烟草K<sup>+</sup>通道<em>NtSKOR</em>基因的克隆及表达分析

doi:10.7668/hbnxb.2018.05.014

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摘要钾通道是植物吸收和转运K⁺的主要蛋白质，SKOR属于Shaker通道外整流家族，在植物低钾胁迫反应中起关键作用。为了研究烟草NtSKOR基因在非生物逆境胁迫中的功能和作用，以普通烟草K326为试验材料，同源克隆一个NtSKOR基因cDNA全长，利用实时荧光定量PCR表征基因表达模式，结合生物信息学分析，对蛋白的理化性质、结构域、磷酸化位点和进化关系等进行初步预测。生物信息学分析表明，该基因全长2 484 bp，编码827个氨基酸残基，预测分子量为94.75 ku，等电点为6.52。该蛋白最大二级结构元件为α-螺旋，最小为β-转角，含有6个跨膜结构域（S1~S6），具有丝氨酸、苏氨酸和酪氨酸3种不同激酶磷酸化位点。进化分析表明，NtSKOR与美花烟草和绒毛状烟草SKOR蛋白同源性高，分别是99%和96%，故命名为NtSKOR。组织表达分析发现，该基因在烟草根、茎、叶、花中均有表达，其中根中表达量最高，花中表达量最低。逆境胁迫试验表明，该基因能快速响应低钾、高盐、干旱、H₂O₂、ABA和4℃等非生物逆境处理。由此推测，NtSKOR在烟草非生物逆境胁迫起着重要调节作用，为今后进一步深入研究NtSKOR功能提供了理论基础。

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关键词 ：烟草； NtSKOR；克隆；钾通道；基因表达

Abstract：Potassium channel is the main protein of plant uptake and translocation K⁺,and SKOR (Stelar K⁺ outwaed rectifier) belongs to Shaker channel family exogenous rectifier,which plays a key role in response to low potassium stress response in plants.In order to study the function and role of tobacco NtSKOR gene in abiotic stress response,one NtSKOR gene was cloned from Nicotiana tabacum K326 with homologous cloning strategy,gene expression patterns were analyzed by Real-time quantitative PCR,and their physicochemical properties,domains,phosphorylation sites and evolutionary relationships of proteins were predicted by bioinformatic analysis. Bioinformatics analysis showed that this gene contained 2 484 bp and encoded 827 amino acid. The predicted molecular weight was 94.75 ku and the isoelectric point (pI) was 6.52. The largest secondary structure element of this protein was α-helix,and the smallest was β-turn. NtSKOR contained six transmembrane domains(S1-S6),with three different kinase phosphorylation sites for serine,threonine and tyrosine. Phylogenetic analysis showed that the protein had 99% homology with Nicotiana sylvestris and 96% homology with Nicotiana tompentosiformis,thus it was named as NtSKOR. Expression patterns showed that the gene was expressed in roots,stems,leaves and flowers in mature stage,which had the highest expression in roots and the lowest expression level in flowers. Expression patterns under abiotic stress indicated the gene could respond rapidly to the adverse conditions of low potassium,high salt,drought,H₂O₂,ABA and 4℃. This suggests that NtSKOR gene plays an important regulatory role in tobacco abiotic stress,and the results provide a theoretical basis for further in-depth study of the NtSKOR function.

Key words： Tobacco ; NtSKOR ; Cloning ; Potassium channel ; Gene expression

收稿日期: 2018-07-18

:	Q78
	S572.03

基金资助:植物生理学与生物化学国家重点实验室开放课题（SKLPPBKF 1505；SKLPPBKF 1506）

通讯作者: 李立芹(1974-),女,山东东阿人,副教授,博士,硕士生导师,主要从事植物分子生物学研究。

作者简介: 卓维(1994-),女,重庆人,在读硕士,主要从事植物分子生物学研究。

引用本文:

卓维, 陈倩, 杨尚谕, 李佳皓, 彭双, 王静, 李立芹. 烟草K⁺通道NtSKOR基因的克隆及表达分析[J]. 华北农学报, 2018, 33(5): 99-105. doi: 10.7668/hbnxb.2018.05.014.
ZHUO Wei, CHEN Qian, YANG Shangyu, LI Jiahao, PENG Shuang, WANG Jing, LI Liqin. Cloning and Expression Analysis of Potassium Channel NtSKOR Gene in Nicotiana tabacum. ACTA AGRICULTURAE BOREALI-SINICA, 2018, 33(5): 99-105. doi: 10.7668/hbnxb.2018.05.014.

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http://www.hbnxb.net/CN/10.7668/hbnxb.2018.05.014 或 http://www.hbnxb.net/CN/Y2018/V33/I5/99

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