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| 陆地棉小GTP结合蛋白基因GhROP1和GhROP8的克隆及表达分析 |
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胡子曜1,武晓玉1,雷建峰2,程贯富1,刘超1,代培红1,梁春燕1,徐诗佳2,李月1*
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| (1.新疆农业大学 生命科学学院, 乌鲁木齐 830052;2.新疆农业大学 农学院/教育部棉花工程研究中心/农业生物技术重点实验室, 乌鲁木齐 830052) |
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| 摘要: |
| 为探究棉花ROP基因在非生物逆境胁迫及外源激素处理下的表达模式，利用同源克隆的方法获得2个陆地棉(Gossypium hirsutum L.)ROP基因GhROP1和GhROP8，通过生物信息学方法分析其理化性质、结构及进化关系，并利用荧光定量PCR(qRT-PCR)技术对其组织表达的特异性及不同逆境胁迫和外源激素处理下的表达模式进行分析。结果表明:1)GhROP1和GhROP8基因的开放阅读框(ORF)分别为591和630 bp，各编码197和210个氨基酸，均含有1个保守的RHO结构域。2)多序列比对显示2个GhROPs蛋白均符合ROP蛋白结构特征且与其他物种ROP蛋白具有高度的相似性，聚类分析表明，GhROP1蛋白属于Ⅰ类ROP蛋白，GhROP8蛋白属于Ⅱ类。3)2个GhROPs基因在不同组织中均有表达且存在组织特异性:GhROP1基因在真叶中表达量最高，在根部和茎部表达量较低；GhROP8基因在根部表达量最高，其他组织中均为低表达量。2个GhROPs基因对于干旱、高盐、低温及高温等非生物逆境胁迫和外源脱落酸(ABA)、生长素(IAA)的处理均有响应:2个GhROPs基因表达均受干旱胁迫诱导；GhROP1基因表达受高盐胁迫抑制，而GhROP8基因受高盐胁迫诱导；低温处理下2个GhROPs基因表达均呈先下调后上调趋势；高温处理下GhROP1基因表达均呈先下调后上调趋势，GhROP8基因表达呈先上调后下调再上调趋势；2个GhROPs基因表达均受外源ABA抑制；外源IAA处理下GhROP1基因呈先上调后下调再上调趋势，GhROP8基因表达受外源IAA诱导。综上，GhROP1和GhROP8是参与棉花响应多种逆境胁迫的重要基因。 |
| 关键词: 棉花 GhROPs 基因克隆 表达分析 逆境胁迫 |
| DOI：10.11841/j.issn.1007-4333.2023.04.02 |
| 投稿时间：2022-06-15 |
| 基金项目:新疆维吾尔自治区重大科技专项(2021A02001-3) |
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| Molecular cloning and expression analysis of small GTP-binding protein genes GhROP1 and GhROP8 in cotton (Gossypium hirsutum L.) |
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HU Ziyao1,WU Xiaoyu1,LEI Jianfeng2,CHENG Guanfu1,LIU Chao1,DAI Peihong1,LIANG Chunyan1,XU Shijia2,LI Yue1*
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| (1.College of Life Sciences, Xinjiang Agricultural University, Urumqi 830052, China;2.College of Agronomy/Research Center of Cotton Engineering of Ministry of Education/Laboratory of Agricultural Biotechnology, Xinjiang Agricultural University, Urumqi 830052, China) |
| Abstract: |
| In order to explore the expression patterns of ROP genes in cotton under different abiotic stresses, GhROP1 and GhROP8 from Gossypium hirsutum L. were obtained by homologous cloning. Their physicochemical properties, structures and evolutionary relationships were analyzed by bioinformatics methods, and tissue expression specificity was explored by fluorescence quantitative PCR(qRT-PCR)technology and their expression patterns under different stress and exogenous hormone treatments were analyzed. The results show that: 1)The open reading frames(ORFs)of GhROP1 and GhROP8 genes are 591 and 630 bp in length, which respectively encodes 197 and 210 amino acids and both containing a conserved RHO domain. 2)Multiple sequence alignment result shows that the two GhROPs are conformed to have the structural characteristics of ROP proteins and has a high degree of homology with ROP proteins of other species. Cluster analysis shows that GhROP1 protein belongs to class I ROP protein, and GhROP8 protein belongs to class II ROP protein; 3)The two GhROPs genes are expressed in different tissues and have certain tissue specificity. The expression level of GhROP1 gene is the highest in leaves, and its expression level was lower in roots and stems. The expression of GhROP8 gene is the highest in roots and low in other tissues. The two genes are sensitive to abiotic stresses such as drought, high salt, low and high temperature and exogenous abscisic acid(ABA), auxin(IAA)all treatments have different degrees of response: The expressions of the two genes are induced by drought stress. The expression of GhROP1 gene is inhibited by high salt stress, while that of GhROP8 gene is induced by high salt stress; The expression of GhROP1 gene is inhibited by high salt stress, while GhROP8 gene is induced by high salt stress; Under low temperature treatment, the expression of two GhROPs genes decreased first and then increased; The expression of GhROP1 gene is down-regulated and then up-regulated under high temperature treatment, and the expression of GhROP8 gene is up-regulated and then down-regulated and then up-regulated; The expression of two GhROPs genes is inhibited by exogenous ABA; Under exogenous IAA treatment, GhROP1 gene is up-regulated first, then down-regulated and up-regulated again, and GhROP8 gene expression is induced by exogenous IAA. In conclusion, GhROP1 and GhROP8 genes may be important regulators of stress resistance in cotton, and this study provides a theoretical basis for further investigation of their biological function in cotton stress response. |
| Key words: cotton GhROPs gene cloning expression analysis stress |
