松材线虫Bx-TIMP克隆及功能研究  

Cloning and Functional Analysis of Bursaphelenchus xylophilus Bx-TIMP

作　　者：杨帆[1,2] 零雅茗[1,2,5] 舒红 姜生伟 王佳楠 李丹蕾 YANG Fan;LING Yaming;SHU Hong;JIANG Shengwei;WANG Jianan;LI Danlei(School of Forestry,Northeast Forestry University,Harbin 150040,China;Key Laboratory of Alien Forest Pest Monitoring and Control-Heilongjiang Province,Northeast Forestry University,Harbin 150040,China;Liaoning Provincial Station of Forest and Grassland Pest Control and Quarantine,Shenyang 110001,China;Liaoning Provincial Key Laboratory of Dangerous Forest Pest Management and Control,Shenyang Institute of Technology,Shenyang 113122,China;Shiwandashan National Nature Reserve Administration,Fangchenggang 535500,China)

机构地区：[1]东北林业大学林学院,哈尔滨150040 [2]黑龙江省外来林木病虫害监测与防控重点实验室(东北林业大学),哈尔滨150040 [3]辽宁省林业和草原局有害生物防治检疫工作站,沈阳110001 [4]沈阳工学院,辽宁省危险性林业有害生物防控重点实验室,沈阳113122 [5]十万大山国家级自然保护区管理局,广西防城港535500

出　　处：《森林工程》2022年第2期14-19,共6页Forest Engineering

基　　金：国家自然科学基金项目(31971656);辽宁省科学技术计划(2019JH2/10200001)。

摘　　要：为探究Bx-TIMP基因在松材线虫(Bursaphelenchus xylophilus)致病过程中的功能,对Bx-TIMP基因进行克隆及分析,并验证基因沉默后松材线虫致病性变化。PCR法克隆Bx-TIMP,应用TMHMM 2.0 server和SignalP 4.1 Server分析该基因编码蛋白质的跨膜结构域和信号肽;应用原位杂交技术确定该基因在松材线虫体内表达部位;应用RNAi技术,分析沉默该基因后松材线虫对红松(Pinus koraiensis)致病性变化。该基因CDS区全长363 bp,编码120个氨基酸,编码蛋白具有跨膜结构域及信号肽。原位杂交表明该基因在松材线虫食道腺中表达。基因沉默后,松材线虫对红松致病性减弱。结果表明,Bx-TIMP为效应因子基因,与松材线虫致病性相关。本研究揭示Bx-TIMP基因是松材线虫危害松树致使其发病的关键基因,为进一步明确松材线虫致病机理提供理论依据,为研发松材线虫的防治技术奠定理论基础。To explore the function of the gene Bx-TIMP during Bursaphelenchus xylophilus infecting Pinus sp.,Bx-TIMP was cloned and analyzed,then the gene was silenced by RNAi and vitrificated.The transmembrane domain and signal peptide of the protein encoded by Bx-TIMP was analyzed by TMHMM 2.0 server and SignalP 4.1 Server.The expression site of the gene in B.ylophilus was determined by in-situ hybridization.By silencing the gene with RNAi,the pathogenicity of B.xylophilus to P.koraiensis was analyzed.The gene had a total length of 363 bp and encoded 120 amino acids.The protein encoded by Bx-TIMP had a transmembrane domain and signal peptide and the results of in-situ hybridization showed that the gene was expressed in the esophageal gland of B.xylophilus,which accorded with the characteristics of effectors.The symptoms of P.koraiensis in Bx-TIMP-RNAi group were significantly weaker than those in the control group.This study showed that Bx-TIMP was an effector gene,which was related to the pathogenicity of B.xylophilus.This study revealed that Bx-TIMP gene is the key gene of B.xylophilus endangering pine trees,which provided a theoretical basis for further clarifying the pathogenesis of B.xylophilus and laying a theoretical foundation for the development of control technology of B.xylophilus.

关 键 词：松材线虫 致病性 效应因子 红松 基因沉默 

分 类 号：S763.18[农业科学—森林保护学]