木香花抗黑斑病基因RbRdr1的克隆与表达分析  

Cloning and expression analysis of black spot resistance gene RbRdr1 in Rosa banksiae

作　　者：范元兰 张真建 陈宇春 蹇洪英[1,3] 晏慧君 张颢[1,3] 周宁宁 王其刚[1,3] 邱显钦 FAN Yuanlan;ZHANG Zhenjian;CHEN Yuchun;JIAN Hongying;YAN Huijun;ZHANG Hao;ZHOU Ningning;WANG Qigang;QIU Xianqin(Flower Research Institute of Yunnan Academy of Agricultural Sciences,Kunming 650205,China;School of Life Sciences,Yunnan University,Kunming 650091,China;National Ornamental Horticultural Engineering Technology Research Center,Kunming 650205,China)

机构地区：[1]云南省农业科学院花卉研究所,昆明650205 [2]云南大学生命科学学院,昆明650091 [3]国家观赏园艺工程技术研究中心,昆明650205

出　　处：《植物生理学报》2020年第12期2716-2724,共9页Plant Physiology Journal

基　　金：国家重点研发计划(2018YFD1000400);云南省科技厅科技人才引进与培养专项“科技领军人才”(2016HA005);云南省中青年学术技术带头人后备人才培养项目(2015-HB078);云南省技术创新人才培养项目(2019HB064);云南万人计划青年拔尖人才和云岭产业技术领军人才培养项目。

摘　　要：月季黑斑病是由真菌蔷薇盘二孢(Marssonina rosae)引起的。本研究以高抗黑斑病菌的野生种木香花(Rosa banksiae)作为实验材料,采用TA克隆法和RACE技术,克隆获得抗黑斑病候选基因RbRdr1 (GenBank登录号MK584935),并对其进行相关生物信息学分析。序列分析结果显示, RbRdr1含有1个3 396 bp的完整的开放阅读框,能编码1 131个氨基酸;其编码的蛋白是一个亲水性蛋白,亚细胞定位于线粒体及过氧化物酶体上,不具有信号肽及跨膜结构域。氨基酸同源序列多重比对发现,该基因与野蔷薇(R.multiflora)和玫瑰(R.rugosa)具有较高的同源性,其中与野蔷薇同源性为86%,玫瑰同源性为84%。实时荧光定量PCR结果表明,该基因在木香花不同组织中均有表达且存在组织特异性,在幼叶及成熟叶中的表达量相对较高,在根、茎及花瓣中的表达量相对较低。RbRdr1基因在受黑斑病菌侵染不同时期的成熟叶片中,其相对表达量均表现为诱导积极上调,且在侵染6 h最高。综上, RbRdr1基因可能参与了木香花与黑斑病菌早期的互作过程。Black spot of roses is caused by the fungus Marssonina rosae. In this study, one wild species with high resistance to Marssonina rosae, Rosa banksiae was used as the experimental material. The candidate gene RbRdr1 from R. banksiae was cloned and obtained based on TA cloning method and RACE technology, and its registration number in GenBank was MK584935. Related bioinformatics analysis was carried out. Sequence analysis showed that RbRdr1 contained a complete open reading frame of 3 396 bp, encoding 1 131 amino acids. The encoded protein was a hydrophilic protein. Its subcellular location was located on mitochondria and peroxidase. And it had no signal peptide and transmembrane domain. Multiple alignment of amino acid homology sequences showed that the gene had high homology with other Rosa species, with 86% homology with R. multiflora and 84% homology with R. rugosa. Real-time fluorescence quantitative PCR results showed that the gene was expressed in different tissues of R. banksiae and had tissue specificity. Its expression level in young leaves and mature leaves were relatively high, while the expression level in roots, stems and petals were relatively low. Its relative expression level in mature leaves inoculated by M. rosae at different stages was up-regulated positively, and it had reached the highest level at 6 h. The results showed that the candidate gene may be involved in the early interaction process between R. banksiae and M. rosae.

关 键 词：木香花 黑斑病菌 RbRdr1基因 RACE技术 生物信息学分析 实时荧光定量PCR 

分 类 号：S436.8[农业科学—农业昆虫与害虫防治] Q943.2[农业科学—植物保护]