棉花转甲基酶基因GhBSMT1的克隆、原核表达及表达谱分析

doi:10.16409/j.cnki.2095-039x.2017.04.005

中国生物防治学报 ›› 2017, Vol. 33 ›› Issue (4): 463-471.DOI: 10.16409/j.cnki.2095-039x.2017.04.005

棉花转甲基酶基因GhBSMT1的克隆、原核表达及表达谱分析

井维霞^1,2, 黄欣蒸², 刘丹凤², 张强², 寇俊凤², 曲爱军¹, 张永军², 郭予元²

1. 山东农业大学植物保护学院, 泰安 271018;
2. 中国农业科学院植物保护研究所/植物病虫害生物学国家重点实验室, 北京 100193

收稿日期:2017-03-18 出版日期:2017-08-08 发布日期:2017-08-11
通讯作者: 曲爱军,副教授,E-mail:aijunqu1965@163.com;张永军,研究员,E-mail:yjzhang@ippcaas.cn
作者简介:井维霞,硕士研究生,E-mail:weixiajing2012@163.com
基金资助:
国家自然科学基金（31471778，31672038，31621064）

Cloning and Prokaryotic Expression of the Methyltransferase Gene GhBSMT1 from Gossypium hirsutum and Its Expression Profile Analysis

JING Weixia^1,2, HUANG Xinzheng², LIU Danfeng², ZHANG Qiang², KOU Junfeng², QU Aijun¹, ZHANG Yongjun², GUO Yuyuan²

1. College of Plant Protection, Shandong Agricultural University, Tai'an 271018, China;
2. State Key Laboratory for Biology of Plant Diseases and Insect Pests/Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China

Received:2017-03-18 Online:2017-08-08 Published:2017-08-11

摘要/Abstract

摘要： 采用RT-PCR结合RACE技术从陆地棉中棉所12中克隆得到全长为1280 bp的苯甲酸/水杨酸羧基甲基转移酶基因序列，命名为GhBSMT1。预测该转移酶分子量40.8 kD，等电点6.12，为不稳定的亲水蛋白。通过原核表达获得大小约41 kD的可溶性重组目标蛋白。利用实时荧光定量PCR技术检测了棉花中GhBSMT1在植物激素处理和棉铃虫取食后表达转录情况，发现GhBSMT1在棉花的根、茎和叶中均有表达。水杨酸甲酯（MeSA）处理能够引起GhBSMT1下调表达，而茉莉酸甲酯（MeJA）处理对靶标基因影响不显著。另外，棉铃虫取食棉株6 h或48 h后，GhBSMT1表达量显著降低。结果表明，GhBSMT1在棉花各组织均有分布并受外源MeSA调控，该基因的表达还被棉铃虫取食抑制。

关键词: 陆地棉, 苯甲酸/水杨酸羧基甲基转移酶, 序列特征, 原核表达, 组织表达谱

Abstract: In this study, a novel benzoic acid/salicylic acid carboxyl methyltransferase gene was cloned from Gossypium hirsutum CCRI 12 by using RT-PCR and RACE-PCR, which contained 1280 bp and named as GhBSMT1. The transferase was predicted to have a molecular weight of 40.8 kD and isoelectric point of 6.12, which was unstable hydrophilic protein. A recombinant soluble protein with a size of about 41 kD was also obtained by prokaryotic expression. The real-time quantitative PCR analysis indicated that GhBSMT1 was constitutively expressed in leaves, stems and roots of cotton plants. The transcript accumulation of GhBSMT1 in leaves declined significantly when the leaves were treated with methyl salicylate (MeSA), whereas methyl jasmonate (MeJA) had no obvious influence on the expression of GhBSMT1. Moreover, the expressions of GhBSMT1 were reduced significantly after 6 h or 48 h feeding by Helicoverpa armigera larvae. In general, the GhBSMT1 is expressed in all tissues of cotton plants and regulated by MeSA and H. armigera infestation.

Key words: Gossypium hirsutum, benzoic acid/salicylic acid carboxyl methyltransferases, sequence characteristics, prokaryotic expression, tissue expression profile

中图分类号:

S435.62

井维霞, 黄欣蒸, 刘丹凤, 张强, 寇俊凤, 曲爱军, 张永军, 郭予元. 棉花转甲基酶基因GhBSMT1的克隆、原核表达及表达谱分析[J]. 中国生物防治学报, 2017, 33(4): 463-471.

JING Weixia, HUANG Xinzheng, LIU Danfeng, ZHANG Qiang, KOU Junfeng, QU Aijun, ZHANG Yongjun, GUO Yuyuan. Cloning and Prokaryotic Expression of the Methyltransferase Gene GhBSMT1 from Gossypium hirsutum and Its Expression Profile Analysis[J]. journal1, 2017, 33(4): 463-471.

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棉花转甲基酶基因GhBSMT1的克隆、原核表达及表达谱分析

Cloning and Prokaryotic Expression of the Methyltransferase Gene GhBSMT1 from Gossypium hirsutum and Its Expression Profile Analysis

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