烟草ERF基因<em>NtRAP2-7</em>的表达载体构建及表达模式分析

doi:10.7668/hbnxb.2018.04.015

摘要/Abstract

摘要： 为了研究烟草NtRAP2-7基因在非生物胁迫响应中的功能，采用同源克隆的方法，在烟草品种K326中克隆到1个ERF转录因子基因NtRAP2-7。并利用生物信息学软件分析了该基因编码蛋白的理化性质、空间结构与系统进化关系。序列分析结果表明，该基因序列全长1 398 bp，共编码465个氨基酸残基，预测蛋白的分子量为51.16 ku。该蛋白包含一个由3个β-折叠及1个α-螺旋组成的保守结构，并含有58个磷酸化位点。亚细胞定位预测表明，该蛋白主要定位于细胞核，并包含单分型的核定位信号序列。进化分析表明，NtRAP2-7蛋白与林烟草RAP2-7蛋白序列同源性最高，为98%。运用qRT-PCR技术进行了该基因的表达模式分析，组织表达分析发现，NtRAP2-7基因在烟草根、茎、叶、花中均有表达，但在根中的表达量最高。该基因的表达在低钾、高盐、干旱、ABA、H₂O₂处理下受到诱导和在低温处理下受到抑制，结果表明，烟草NtRAP2-7基因参与了烟草的非生物胁迫应答反应。并利用双酶切法成功构建了pBI121-NtRAP2-7过表达载体，为今后该基因在非生物胁迫下的功能研究奠定了基础。

关键词: 烟草, NtRAP2-7基因, 基因表达, 非生物胁迫

Abstract: In order to study the function of tobacco NtRAP2-7 gene in abiotic stress response, NtRAP2-7 was cloned from K326 by homologous cloning. And the bioinformatics software was used to analyze the physicochemical properties, spatial structure and phylogenetic analysis of the encoded protein. Sequence analysis results showed that the gene contained 1 398 bp in length which encoded 465 amino acid residues and the predicted molecular weight of this protein was 51.16 ku. Based on 3-D modeling and secondary structure analysis, the protein consisted of three β-sheet and one α-helix and contained 58 phosphorylation sites. The subcellular localization prediction showed that the protein was mainly located in the nucleus and contained a monopartite nuclear location signal sequence. Phylogenetic analysis showed that NtRAP2-7 protein had the highest homology with Nicotiana sylvestris RAP2-7 protein sequence, which was 98%.The expression pattern analysis of the gene was carried out by qRT-PCR. Tissue expression analysis showed that the gene was expressed in the roots, stems, leaves and flowers, but had the highest expression level in roots. The expression levels of NtRAP2-7 was affected by abiotic stress treatments, indicating that the NtRAP2-7 was involved in the tobacco abiotic stress response. The pBI121-NtRAP2-7 overexpression vector was successfully constructed by double enzyme digestion, which laid the foundation for further study on the function of this gene in abiotic stress.

Key words: Tobacco, NtRAP2-7, Gene expression, Abiotic stress

中图分类号:

Q78
S572.03

陈倩, 卓维, 罗静, 杨尚谕, 鲁黎明, 李立芹. 烟草ERF基因NtRAP2-7的表达载体构建及表达模式分析[J]. 华北农学报, 2018, 33(4): 104-111. doi: 10.7668/hbnxb.2018.04.015.

CHEN Qian, ZHUO Wei, LUO Jing, YANG Shangyu, LU Liming, LI Liqin. Construction of Expression Vector and Expression Analysis of Tobacco ERF Gene NtRAP2-7[J]. Acta Agriculturae Boreali-Sinica, 2018, 33(4): 104-111. doi: 10.7668/hbnxb.2018.04.015.

http://www.hbnxb.net/CN/Y2018/V33/I4/104

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烟草ERF基因NtRAP2-7的表达载体构建及表达模式分析

Construction of Expression Vector and Expression Analysis of Tobacco ERF Gene NtRAP2-7

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烟草ERF基因NtRAP2-7的表达载体构建及表达模式分析

Construction of Expression Vector and Expression Analysis of Tobacco ERF Gene NtRAP2-7

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