普通菜豆生长素调节蛋白基因PvARP1的克隆及表达分析

doi:10.15933/j.cnki.1004-3268.2019.09.012

摘要/Abstract

摘要： 生长素调节蛋白（Auxin regulating protein，ARP）是参与植物生长和发育调控的重要因子，也是植物抗病反应中具有重要作用的蛋白质分子。为了解该蛋白质基因表达特性，利用普通菜豆（Phaseolus vulgaris L.)表达序列标签（EST）克隆了编码普通菜豆ARP蛋白的cDNA序列并进行相关分析。序列分析表明，cDNA片段长2 428 bp，具有1个1 818 bp的开放阅读框，GenBank登录号为MK301448，将其命名为PvARP1。该基因编码605个氨基酸，预测蛋白质分子质量为68.26 ku，编码的蛋白质不含跨膜区、无信号肽。同源分析结果显示，PvARP1基因编码蛋白质与小豆（Vigna angularis）ARP蛋白亲缘关系最近，达到94%。荧光定量PCR分析结果表明，PvARP1基因受镰孢菌菜豆专化型FOP-DM01菌株诱导表达，接种病原菌24 h抗病材料260205根中PvARP1基因的表达量达到最高，提升至其接种0 h表达量的111倍，不同接种时间260205根中PvARP1基因的表达量均高于感病材料BRB130，而且PvARP1基因受吲哚-3-乙酸诱导表达量显著或极显著提高。此外，菜豆花和荚中PvARP1基因表达量明显高于根、茎和叶。vARP1基因在大肠杆菌中可诱导表达为68 ku的重组蛋白，体外具有酰胺合成酶活性，可能参与调节植物细胞内吲哚-3-乙酸水平。表明PvARP1基因可能通过吲哚-3-乙酸介导的信号途径参与菜豆对FOP-DM01菌株的防御反应，推测菜豆PvARP1基因与镰孢菌枯萎病的抗病性有关。

关键词: 普通菜豆, 镰孢菌枯萎病, 生长素调节蛋白, 基因表达分析, 抗病反应

Abstract: Auxin-regulating protein (AＲP) is an important factor involved in the regulation of plant growth and development，and is also a protein molecule that plays an important role in plant disease resist ance． In order to understand the gene expression characteristics of this protein，the cDNA sequence enco ding the AＲP protein was cloned and analyzed by using the expression sequence tag(EST) of common bean(Phaseolus vulgaris L.) .Sequence analysis indicated that the cDNA fragment was 2 428 bp in length and named as PvAＲP1.It contained an open reading frame(OＲF) of 1 818 bp，and the accession number was MK301448.The gene encoded 605 amino acids and the predicted protein molecular weight was 68.26 ku.The encoded protein did not contain transmembrane region and signal peptide.The results of homolo gous analysis showed that the close relationship between PvAＲP1 protein and AＲP protein of Vigna angu laris reached 94%.eal time PCＲ analysis showed that PvAＲP1 gene was regulated by Fusarium oxyspo rum f.sp.phaseoli FOP-DM01 isolate.The expression of PvAＲP1 gene at 24 h after inoculation was in creased to 111 times of that at 0 h as control in the roots of the resistant material 260205 to the disease， and the expression level in 260205 roots was higher than the susceptible material BＲB130 at all time points.The expression of PvAＲP1 gene was significantly enhanced by indole-3-acetic acid(IAA) .In addi tion，the expression level in the flowers and pods was obviously higher than the roots，stems and leaves.The PvAＲP1 gene could be expressed as a 68 ku protein in E.coli BL21(DE3) ，which had the amide synthetase activity in vitro，and might regulate the active IAA level in plant cells.This study indicated that the PvAＲP1 gene probably participated in the defense response of common bean to the FOP-DM01 isolate through the IAA-mediated signaling pathway.It is speculated that the PvAＲP1 gene is related to the resist ance of fusarium wilt in common bean，and has the great significance for directing the molecular breeding of resistance in common bean．

Key words: Common bean, Fusarium wilt, Auxin-regulating protein, Gene expression analysis, Ｒesist ant response

中图分类号:

S643.1

薛仁风, 丰明, 赵阳, 陈剑, 李韬, 葛维德. 普通菜豆生长素调节蛋白基因PvARP1的克隆及表达分析[J]. 河南农业科学, 2019, 48(9): 82-89.

XUE Ｒenfeng, FENG Ming, ZHAO Yang, CHEN Jian, LI Tao, GE Weide. Cloning and Expression Analysis of PvAＲP1 Gene Encoding Auxin-regulating Protein in Common Bean[J]. Journal of Henan Agricultural Sciences, 2019, 48(9): 82-89.

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