利用小RNA深度测序技术鉴定白芷病毒病病原  

Identification of the pathogens of Angelica dahurica virus using small RNA deep sequencing technology

作　　者：杜江[1] 姚东作 王晨燕 崔丽艳 王德富[1] 牛颜冰[1] Du Jiang;Yao Dongzuo;Wang Chenyan;Cui Liyan;Wang Defu;Niu Yanbing(College of Life Sciences,Shanxi Agricultural University,Jinzhong 030801,China;College of Grassland Sciences,Shanxi Agricultural University,Jinzhong 030801,China)

机构地区：[1]山西农业大学生命科学学院,山西晋中030801 [2]山西农业大学草业学院,山西晋中030801

出　　处：《山西农业大学学报（自然科学版）》2023年第1期75-81,共7页Journal of Shanxi Agricultural University(Natural Science Edition)

基　　金：山西省基础研究计划(20210302124676);山西农业大学科技创新基金项目(2020BQ51);财政部和农业农村部:国家现代农业产业技术体系(CARS-21)。

摘　　要：[目的]白芷(Angelica dahurica)是一种药用历史悠久的传统中药,本研究利用小RNA深度测序技术明确引起山西太谷白芷花叶、褪绿症状的病原,为白芷病毒病的防控提供依据。[方法]将采集的白芷病样进行小RNA深度测序,通过RT-PCR和序列相似性分析明确白芷病毒病的分类地位。[结果]白芷样品经小RNA深度测序共获得22158005个原始序列,将拼接contigs与NCBI中的病毒数据库进行比对注释,结果显示比对到黄瓜花叶病毒(Cucumber mosaic virus,CMV)3条正义链RNA 3、RNA 2、RNA 1的比对率分别为2.14%、1.28%和1.10%。利用特异引物克隆了CMV-BZ RNA3的外壳蛋白和移动蛋白基因全序列。通过序列比对分析,发现CMV-BZ CP氨基酸序列和核苷酸序列分别与CMV亚组ⅠB中CMV分离物(CMV-SD-YT和CMV-SXCH)和(CMV-SD-QD)的相似性最高,分别为99.1%和99.5%;MP氨基酸序列和核苷酸序列分别与CMV亚组Ⅰ分离物Ah的相似性最高,为99.3%和99.0%。MP和CP氨基酸序列系统进化分析表明,CMV-BZ与中国大多数CMV分离物聚为一类,属于CMV亚组Ⅰ中的成员。[结论]首次在山西白芷上检测到CMV,该研究有助于深入了解CMV的分子进化,为白芷产业的健康发展提供一定启示,也为病毒病的防治提供一定的理论依据。[Objective]Angelica dahurica is a traditional Chinese medicinal plant with a long history of use.In this study,small RNA deep sequencing technology was used to identify the pathogen causing leaf yellowing and flower blight symptoms in An⁃gelica dahurica in Taigu,Shanxi Province,providing a basis for the prevention and control of CMV.[Methods]A.dahurica samples were collected and subjected to small RNA deep sequencing,and the classification status of A.dahurica CMV isolates(CMV⁃BZ)were determined by RT⁃PCR and sequence similarity analysis.[Results]A total of 22158005 original sequences were obtained from A.dahurica samples by small RNA deep sequencing.The spliced contigs were compared and annotated with the virus database in NCBI,and the results showed that the alignment rates of three positive⁃sense RNA segments(RNA3,RNA2,and RNA1)of Cucumber mosaic virus(CMV)were 2.14%,1.28%and 1.10%,respectively.The complete sequences of coat protein and movement protein genes of CMV⁃BZ RNA3 were cloned using specific primers.Sequence align⁃ment analysis showed that the amino acid and nucleotide sequences of CMV⁃BZ CP were most similar to those of CMV isolates(CMV⁃SD⁃YT and CMV⁃SXCH)and(CMV⁃SD⁃QD)in CMV subgroup IIB,with similarities of 99.1%and 99.5%,respec⁃tively;the amino acid and nucleotide sequences of CMV⁃BZ MP were most similar to those of the CMV subgroup I isolate Ah,with similarities of 99.3%and 99.0%,respectively.Phylogenetic analysis of MP and CP amino acid sequences showed that CMV⁃BZ clustered with most CMV isolates in China and belonged to the members of CMV subgroup I.[Conclusion]To the best of our knowledge,this is the first time that CMV was detected on A.dahurica in Shanxi province,providing insight into the molecular evolution of CMV and a theoretical basis for the prevention and control of virus diseases,as well as contributing to the healthy development of A.dahuricae industry.

关 键 词：白芷 黄瓜花叶病毒 小RNA深度测序 

分 类 号：S432.41[农业科学—植物病理学]