不结球白菜抗病相关基因 BcCPR 1 的克隆和功能分析  

Cloning and function alanalysis of BcCPR 1 gene involved in disease resistance in non-heading Chinese cabbage

作　　者：邵岑 袁水林 王远 朱蔚曈 侯喜林[1] 李英[1] SHAO Cen;YUAN Shuilin;WANG Yuan;ZHU Weitong;HOU Xilin;LI Ying(State Key Laboratory of Crop Genetics and Germplasm Enhancement/Key Laboratory of Biology and Germplasm Enhancement of Horticulture Crop in East China,Ministry of Agriculture and Rural Affairs/College of Horticulture,Nanjing Agricultural University,Nanjing 210095,China)

机构地区：[1]南京农业大学作物遗传与种质创新国家重点实验室/农业农村部华东地区园艺作物生物学与种质创新重点实验室/园艺学院,江苏南京210095

出　　处：《南京农业大学学报》2021年第5期841-849,共9页Journal of Nanjing Agricultural University

基　　金：国家重点研发计划项目(2018YFD1000805);江苏高校优势学科建设工程资助项目(PAPD)。

摘　　要：[目的]本文旨在探究BcCPR 1基因对灰霉菌的抗性及对霜霉菌和非生物胁迫的响应。[方法]采用同源克隆方法从不结球白菜中克隆BcCPR 1基因CDS序列;利用MEGA 7软件对其进行同源性分析;利用农杆菌介导法在烟草中进行亚细胞定位研究及转基因拟南芥植株的获取;利用RT-qPCR技术,对BcCPR 1基因在生物及非生物胁迫处理下的表达水平进行分析。[结果]BcCPR 1基因含有1个1221 bp的开放阅读框(ORF),编码406个氨基酸,与同属的甘蓝和芜菁进化关系最相近,分别为97.54%和92.66%。亚细胞定位结果显示,BcCPR1蛋白定位在细胞膜上。霜霉菌、脱落酸(ABA)、茉莉酸甲酯(MeJA)、水杨酸(SA)和盐处理BcCPR 1基因均有响应,并存在时间表达差异。过表达BcCPR 1基因的转基因拟南芥对灰霉菌抗性增强,同时茉莉酸(JA)途径标记基因PDF 1.2表达量显著上升。[结论]BcCPR1蛋白定位于细胞膜,主要通过茉莉酸(JA)途径调节过表达拟南芥对灰霉菌的抗性,对霜霉菌及非生物胁迫均有响应。[Objectives]This paper aimed to study the resistance of BcCPR 1 gene to Botrytis cinerea and its expression under Hyaloperonospora parasitica and abiotic stresses.[Methods]The CDS sequence of BcCPR 1 gene was cloned by homologous cloning method using non-heading Chinese cabbage as the material.Homology analysis was conducted using MEGA 7 software.By the Agrobacterium-mediated method,the sub-cellular localization was researched in tobacco leaves and transgenic Arabidopsis thaliana was acquired.The expression level of BcCPR 1 was analyzed under biotic and abiotic stress treatments using RT-qPCR.[Results]BcCPR 1 gene contained a 1221 bp open reading frame(ORF),encoding 406 amino acids.Based on the close relationship,the homology between Brassica oleracea var.oleracea and B.rapa was up to 97.54%and 92.66%,respectively.Subcellular localization result showed that the localization of BcCPR1 was on the cell membrane.BcCPR 1 gene responded to the treatments of H.parasitica,abscisic acid(ABA),methyl jasmonate(MeJA),salicylic acid(SA)and salt stress,and there was a time difference in expression.BcCPR 1-overexpressed plants of A.thaliana were more resistant than wild-type plants,while the expression of the jasmonate(JA)pathway marker gene PDF 1.2 increased.[Conclusions]BcCPR1 protein was localized in the cell membrane,regulated the resistance to B.cinerea through JA pathways in overexpression plant of A.thaliana,and responded to H.parasitica and abiotic stresses.

关 键 词：不结球白菜 BcCPR 1 亚细胞定位 灰霉菌 霜霉菌 非生物胁迫响应 

分 类 号：S634.3[农业科学—蔬菜学]