| 赵巍巍1, 杨家强2, 周小毛1*, 吴青君2*.番茄斑萎病毒4种检测方法比较[J].植物保护,2015,41(2):108-113. |
| 番茄斑萎病毒4种检测方法比较 |
| Comparison of four methods for detecting Tomato spotted wilt virus |
| 投稿时间：2014-07-04 修订日期：2014-10-09 |
| DOI： |
| 中文关键词: 番茄斑萎病毒 DAS-ELISA 常规PCR 快速检测试纸条 qRT-PCR 灵敏度 |
| 英文关键词:Tomato spotted wilt virus DAS-ELISA conventional PCR rapid detection test strip qRT-PCR sensitivity |
| 基金项目:国家科技支撑计划(2012BAD19B06); 北京市自然科学基金(6122028); 现代农业产业技术体系北京市叶类蔬菜创新团队建设专项资金(blvt-15); 北京市科委课题(Z121100001212006); 蔬菜有害生物控制与优质栽培北京市重点实验室 |
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| 中文摘要: |
| 本研究根据番茄斑萎病毒(TSWV) S RNA上的核衣壳蛋白(N)基因保守序列设计特异性引物, 比较了4种检测方法的灵敏度。结果表明, 特异性引物可扩增出397 bp的片段, 序列和已发表的TSWV核苷酸序列同源性高达99%, 可用于常规PCR和荧光定量RT-PCR (qRT-PCR)检测。qRT-PCR的灵敏度比快速检测试纸条、双抗体夹心酶联免疫吸附法(DAS-ELISA)和常规PCR分别高出15 625倍、3 125倍和125倍, 且能够准确定量; 常规PCR 灵敏度较高, 但不能准确定量; DAS-ELISA方法适用于批量定性测定, 但检测时间较长; 试纸条法检测速度最快, 但灵敏度最低, 使用时可根据症状程度和试验条件选择适宜的检测方法。 |
| 英文摘要: |
| Specific primers were designed according to the conserved sequence of the nucleocapsid protein (N) gene of Tomato spotted wilt virus (TSWV) S RNA, and the sensitivities of four methods for detecting TSWV were compared. The results showed that a 397 bp fragment was amplified with the specific primers, and the sequence showed a homology of 99% with the published TSWV nucleotide sequence. The designed primers could be used for conventional PCR and quantitative reverse transcription-PCR (qRT-PCR) methods. The sensitivity of qRT-PCR for detecting TSWV was 15 625 fold, 3 125 fold and 125 fold higher than that of the rapid detection test strip (RDTT), double sandwich enzyme-linked immunosorbent assay (DAS-ELISA) and conventional PCR, respectively. Moreover, qRT-PCR method can be accurately quantified. Conventional PCR showed a higher sensitivity, but it cannot be accurately quantified. DAS-ELISA is suitable for batch qualitative detection but it needs a longer detection time. RDTT is the fastest method, but the sensitivity is the lowest. Suitable detection methods can be selected according to the symptom degree and the experimental conditions. |
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