[1]蔡伟,金晶,沈建国,等.TaqMan-MGB 荧光定量IC /TC-ＲT-PCＲ 检测菜豆荚斑驳病毒[J].大豆科学,2016,35(03):489-493.[doi:10.11861/j.issn.1000-9841.2016.03.0489]
　CAI Wei,JIN Jing,SHEN Jian-guo,et al.Detection of Bean Pod Mottle Virus by TaqMan-MGB Ｒeal-time IC/TC-ＲT-PCＲ[J].Soybean Science,2016,35(03):489-493.[doi:10.11861/j.issn.1000-9841.2016.03.0489]
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TaqMan-MGB 荧光定量IC /TC-ＲT-PCＲ 检测菜豆荚斑驳病毒

《大豆科学》[ISSN:1000-9841/CN:23-1227/S] 卷: 第35卷 期数: 2016年03期 页码: 489-493 栏目: 出版日期: 2016-05-20
Title:
Detection of Bean Pod Mottle Virus by TaqMan-MGB Ｒeal-time IC/TC-ＲT-PCＲ
作者:
蔡伟1; 2; 金晶1; 2; 沈建国1; 高芳銮2; 廖富荣3; 吴祖建2
1. 福清出入境检验检疫局/福建省检验检疫技术研究重点实验室，福建福州350300;?
2. 福建农林大学植物病毒研究所，福建福州350002;
3. 厦门出入境检验检疫局检验检疫技术中心，福建厦门361012
Author(s):
CAI Wei1; 2; JIN Jing1; 2; SHEN Jian-guo1; GAO Fang-luan2; LIAO Fu-rong3; WU Zu-jian2
1. Fuqing Entry-Exit Inspection and Quarantine Bureau /Fujian Key Laboratory for Technology Ｒesearch of Inspection and Quaranting，Fuzhou 350300，China;?
2. Institution of Plant Virology，Fujian Agriculture and Forestry University，Fuzhou 350002，China;
3. Xiamen Key Laboratory for Technology Ｒesearchof Inspection and Quarantine，Xiamen 361012，China
关键词:
TaqMan-MGB 荧光定量IC-ＲT-PCＲ;  TaqMan-MGB 荧光定量TC-ＲT-PCＲ;  菜豆荚斑驳病毒;  检测
Keywords:
TaqMan-MGB real-time IC-ＲT-PCＲ;  TaqMan-MGB real-time TC-ＲT-PCＲ;  Bean pod mottle virus ( BPMV) ;  Detection
分类号:
S565. 1
DOI:
10.11861/j.issn.1000-9841.2016.03.0489
文献标志码:
A
摘要:
菜豆荚斑驳病毒( Bean pod mottle virus，BPMV) 是我国重要的检疫性有害生物，BPMV 传入的风险随大豆进境数量增多而加大。本文针对菜豆荚斑驳病毒，以大豆种子提取液为材料，分别建立了TaqMan-MGB 荧光定量IC-ＲTPCＲ和TaqMan-MGB 荧光定量TC-ＲT-PCＲ 快速检测方法。根据GenBank 公布的BPMV 外壳蛋白基因序列，选择其保守区域，设计1 对特异性引物和1 条TaqMan-MGB 探针，测定了TaqMan-MGB 荧光定量IC-ＲT-PCＲ 和TaqMan-MGB 荧光定量TC-ＲT-PCＲ 方法的特异性和灵敏度，并将TaqMan-MGB 荧光定量IC-ＲT-PCＲ、TaqMan-MGB 荧光定量TC-ＲTPCＲ、IC-ＲT-PCＲ 和TC-ＲT-PCＲ 4 种检测方法的灵敏度进行比较。结果表明: 所建立的两种检测方法特异性良好; TCＲT-PCＲ 的灵敏度为10 － 1 倍病毒提取液原液; IC-ＲT-PCＲ 和TaqMan-MGB 荧光定量TC-ＲT-PCＲ 灵敏度相当，为10 － 3倍病毒提取液原液; TaqMan-MGB 荧光定量IC-ＲT-PCＲ 灵敏度最高，为10 － 5 倍病毒提取液原液，是IC-ＲT-PCＲ 和Taq-Man-MGB 荧光定量TC-ＲT-PCＲ 检测方法的102 倍，是TC-ＲT-PCＲ 检测方法的104 倍; 两种检测方法均能较好应用于实际大豆样品的检测。本文所建立的TaqMan-MGB 荧光定量IC/TC-ＲT-PCＲ 检测方法利用抗原特异性或试管非特异性捕捉病毒粒子，无需提取ＲNA，为进境大豆种子上BPMV 的检测提供了稳定、快速、简便的技术依据，其较高的灵敏度和特异性具有较好的应用价值。
Abstract:
Bean pod mottle virus( BPMV) is regarded as an important quarantine plant pathogen in our country，the risk ofBPMV spread from outside increases under the huge soybean import. TaqMan-MGB real-time IC-ＲT-PCＲ and TaqMan-MGBreal-time TC-ＲT-PCＲ methods were developed to detect BPMV by using soybean seed extracts as materials. Based on the conservedregion of coat protein sequences of BPMV in GenBank，primers and TaqMan-MGB probe were designed. The specificityand sensitivity of these two methods were measured and the comparation of sensitivity of TaqMan-MGB real-time IC-ＲT-PCＲ，TaqMan-MGB real-time TC-ＲT-PCＲ，IC-ＲT-PCＲ and TC-ＲT-PCＲ were operated. Two established methods had a high specificity，the comparison results indicated that TC-ＲT-PCＲ shared 10 － 1 crude extracts，IC-ＲT-PCＲ and TaqMan-MGB real-timeTC-ＲT-PCＲ possessed the same sensitivity reaching 10 － 3 crude extracts while TaqMan-MGB real-time IC-ＲT-PCＲ had thehighest sensitivity about 10 － 5 crude extracts. The sensitivity of TaqMan-MGB real-time IC-ＲT-PCＲ was 102 times more thanTC-ＲT-PCＲ，104 times more than TC-ＲT-PCＲ. It was practical to use the two established methods in sample detection. Sothe established TaqMan-MGB real-time IC/TC-ＲT-PCＲ method could meet the need of detection with specific capture of antibodyor nonspecific capture of tube to catch virion and no need to isdate ＲNA. It provides the basic approach to detect BPMVin imported soybean seeds with required specificity and sensitivity characteristics.

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备注/Memo

基金项目: 福州市科技计划项目( 2013-N-54 ) ; 福建出入境检验检疫局科技项目( FK2011-07 ) ; 福建省杰出青年科学基金资助项目( 2014J06008) ; 质检公益性行业科研专项项目( 201410076) 。

第一作者简介: 蔡伟( 1983-) ，男，硕士，农艺师，主要从事植物病毒学及分子生物学研究。E-mail: caiweifjciq@126. com。
通讯作者: 沈建国( 1978-) ，男，博士，高级农艺师，主要从事植物病毒检测及防治研究。E-mail: shenjg_agri@163. com;吴祖建( 1967-) ，男，博士，研究员，主要从事植物病毒学研究。E-mail: wuzujian@126. com。

更新日期/Last Update: 2016-07-24