猕猴桃病程相关蛋白PR-1基因的克隆和功能分析  

Cloning and function analysis of PR-1 gene in Actinidia

作　　者：张敏 宋雅林[1] 林苗苗 王然[1] 李玉阔[1,2] 孙艳香 方金豹[1,2] 苏彦苹[3] 孙雷明 齐秀娟[1,2] ZHANG Min;SONG Yalin;LIN Miaomiao;WANG Ran;LI Yukuo;SUN Yanxiang;FANG Jinbao;SUN Yanping;SUN Leiming;QI Xiujuan(National Key Laboratory for Germplasm Innovation&Utilization of Horticultural Crops/Zhengzhou Fruit Research Institute,Chinese Academy of Agricultural Sciences,Zhengzhou 450009,Henan,China;Zhongyuan Research Center,Chinese Academy of Agricultural Sciences,Xinxiang 453500,Henan,China;Langfang Normal University,Langfang 065000,Hebei,China)

机构地区：[1]果蔬园艺作物种质创新与利用全国重点实验室·中国农业科学院郑州果树研究所,郑州450009 [2]中国农业科学院中原研究中心,河南新乡453500 [3]廊坊师范学院,河北廊坊065000

出　　处：《果树学报》2024年第8期1524-1533,共10页Journal of Fruit Science

基　　金：河南省重大科技专项(221100110400);国家重点研发计划(2022YFD1600700);国家现代农业产业技术体系(CARS-26);中国农业科学院科技创新工程(CAAS-ASTIP-2024-ZFRI-03);河南省现代农业产业技术体系(HARS-22-09-S);国家园艺种质资源库郑州猕猴桃分库(NHGRC2024-NH00-4);河北省重点研发计划项目(20326335D)。

摘　　要：【目的】探究猕猴桃病程相关蛋白(pathogenesis-related proteins,PRs)PR-1基因在响应丁香假单胞杆菌中的功能。【方法】以毛花猕猴桃(Actinidia eriantha)为材料,克隆得到PR-1同源基因AePR-1全长序列,并对其序列进行生物信息学分析。采用实时荧光定量方法检测AePR-1基因在不同组织、花器官以及接种细菌性溃疡病菌(Psa)和不同激素(SA、ABA、GA_(3))处理条件下的表达情况。利用亚细胞定位技术分析AePR-1基因在细胞中的表达位置。通过在本氏烟草中过表达AePR-1基因,验证其在溃疡病菌响应过程中的功能。【结果】猕猴桃AePR-1基因序列全长522 bp,编码173个氨基酸,序列中含有6个保守的半胱氨酸结构基序和4个allergen V5/Tpx-1 related保守结构域。亚细胞定位发现AePR-1定位在细胞膜和细胞质中。AePR-1在猕猴桃根和雌蕊中高表达,且能够响应溃疡病菌及激素处理。过表达AePR-1的烟草在接种溃疡病菌后,叶片病斑数明显少于对照组。【结论】AePR-1基因在溃疡病菌和激素诱导下显著表达且过表达能够增强烟草对溃疡病的抗性,说明猕猴桃PR-1基因在响应生物和非生物胁迫中具有重要作用。【Objective】China is the origin of the kiwifruit(Actinidia spp.),with rich germplasm resources and wide geographical distribution.It is one of the most recently domesticated fruit plants and has become an important horticultural crop.There are 54 species and 21 varieties of the A.Lindl.in the world.Kiwifruit bacterial canker is a devastating disease in kiwifruit industry globally and caused by pathogen Pseudomonas syringae pv.actinidiae(Psa).Psa is highly virulent,and once systemic invade plant may quickly lead plant to death.It has been documented that the Pathogenesis-related 1 protein(PR-1)could resist the spread of viruses,limit the invasion of pathogens and protect plants from adversity stress.In many plant species such as Arabidopsis and tobacco,the overexpression of the PR-1 gene could enhance plant resistance to P.syringae.However,the PR-1 gene in kiwifruit and its role in responses to abiotic stress remain largely unknown.The objective of this study was to explore the function of kiwifruit Pathogenesis-related 1 gene(PR-1)in response to biological stress.This analysis could contribute to in-depth understanding the function of the PR-1 gene in kiwifruit disease resistance.【Methods】Annual grafted treess of kiwifruit species A.eriantha were used as experimental materials.The full-length sequence of the PR-1 homologous gene AePR-1 of A.eriantha was cloned and analyzed by multiple bioinformatic tools.The DNAMAN software was used to compare and analyze the sequence of the AePR-1 gene.The conserved domain of AePR-1 protein sequence was analyzed by NCBI website.The Expasy ProtParam tool was used to predict the molecular weight,theoretical pI,instability index and grand average of hydropathicity(GRAVY)of AePR-1 protein.The Cell-PLoc 2.0 software was used to predict the subcellular localization of PR-1 protein.The phylogenetic relationship between the AePR-1 protein and PR-1 of other plants was analyzed by the MEGA 11.013 software using neighbor-joining method.The qRT-PCR was performed to analyze the expres

关 键 词：猕猴桃 PR-1基因 细菌性溃疡病菌(Psa) 抗病 

分 类 号：S663.4[农业科学—果树学]