作者：李茗乐, 雷荣, 杨藜, 孙夕雯, 杨益芬, 王新一, 范在丰, 段维军, 吴品珊
单位：1 中国农业大学植物保护学院 农业农村部植物检疫性有害生物监测防控重点实验室，北京 100093,2 中国检验检疫科学研究院，北京 100176,3 宁波检验检疫科学技术研究院，浙江 宁波 315012,4 成都海关技术中心，四川 成都 610041,5 大连大学，辽宁 大连 116622
关键词：苜蓿轮枝菌,重组酶聚合酶等温扩增,CRISPR-Cas12a检测系统,快速检测
基金：国家重点研发计划(2021YFC2600400),国家重点研发计划(2021YFC2600402),中国检验检疫科学研究院基本科研业务费项目(2022JK20)
苜蓿轮枝菌Verticillium alfalfae是我国重要的进境植物检疫对象，能够对十几种作物造成严重危害。为了快速检测苜蓿轮枝菌，本研究选用苜蓿轮枝菌的特征序列扩增区域[sequence characterized amplified region (SCAR) marker]作为目标基因，设计了特异性引物Va-RPA-F/Va-RPA-R和CRISPR 引导RNA (Va-crRNA)，建立了重组酶聚合酶等温扩增技术结合CRISPR-Cas12a的快速检测方法。对该检测体系的反应时间、组分浓度等进行优化，结果显示苜蓿轮枝菌基因组DNA在39 ℃恒温条件下扩增30 min后，其扩增产物用基于CRISPR-Cas12a的荧光法和侧向流层析试纸条两种方法检测，可检测到10.6 pg基因组DNA，其灵敏度与实时荧光定量PCR结果相当，加标回收实验结果证明本研究建立的方法可成功用于植物组织中苜蓿轮枝菌的快速检测。"/>template{display:none;}.mag-rich-xref-fn { cursor: pointer;}//长视频$(document).ready(function(){if($("#showLongArticleVideo") && $("#showLongArticleVideo").length && $("#showLongArticleVideo").length>0){mag_ajax_update({ele_id:'showLongArticleVideo',url:mag_currentQikanUrl() + "/CN/article/showLongArticleVideo.do?id="+$("#articleId").val()});}});//可视化图谱$(document).ready(function(){if($("#showArticle_knowledgeMap") && $("#showArticle_knowledgeMap").length && $("#showArticle_knowledgeMap").length>0){jQuery.getJSON("/Jwxb/CN/lexeme/existsCnctstInArticle.do?articleID="+$("#articleId").val(), null, function(json){if(json.result){$("#showArticle_knowledgeMap").show();}});}});$(document).ready(function(){var metaDataStr = $.trim($("#article_metaJson").val());if(metaDataStr!=''){eval("window.metaData="+metaDataStr);}if(window.metaData && (!window.metaData.authorNotesCommon_cn && !window.metaData.authorNotesCorresp_cn && window.metaData.authorNotes_cn)){window.metaData.authorNotesCommon_cn = window.metaData.authorNotes_cn;}if(window.metaData && (!window.metaData.authorNotesCommon_en && !window.metaData.authorNotesCorresp_en && window.metaData.authorNotes_en)){window.metaData.authorNotesCommon_en = window.metaData.authorNotes_en;}var _nlmdtdXml = $("#article_nlmdtdXml").val();var magDir = _nlmdtdXml.replace(/\/[^\/]+$/,'');window.metaData.magDir = magDir;new mag_vue({ el: '#metaVue', //dataUrl: mag_currentQikanUrl() + '/EN/article/getRichHtmlJson.do?articleId='+$("#articleId").val(), data:window.metaData, loading:true, doTextFun:function(text){ return doDataJsonText(text); 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来源：2024年第9期
《菌物学报》期刊编辑部
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