One-step reverse transcription loop mediated isothermal amplification assay for sensitive and rapid detection of Cucurbit chlorotic yellows virus
One-step reverse transcription loop mediated isothermal amplification assay for sensitive and rapid detection of Cucurbit chlorotic yellows virus
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一步逆转录环介导的等温扩增试验灵敏快速检测葫芦褪绿黄化病毒
DOI：
10.1016/j.jviromet.2013.08.037
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发表时间：
2014-01-01
影响因子：
3.1
通讯作者：
Shi, Yan
中科院分区：
文献类型：
--
作者：
Wang, Zhenyue;Gu, Qinsheng;Shi, Yan
开发了一种逆转录环介导的等温扩增 (RT-LAMP) 检测方法来检测葫芦褪绿黄化病毒 (CCYV)。在此过程中，使用从CCYV感染的瓜叶组织中提取的总RNA，合成了与CCYV外壳蛋白基因区域中总共6个序列相匹配的一组4个引物，用于RT-LAMP测定，并确定了最佳反应温度和测定时间。敏感性测定表明，在稀释度为1 x 10(-11)的RT-LAMP反应中可检测到病毒，其敏感性比RT-PCR测定高10(5)倍。 CCYV 和甘薯褪绿矮化病毒 (SPCSV) 的 RT-LAMP 检测对 CCYV 表现出高度特异性。这种简单而灵敏的方法有可能检测现场收集的样品中的 CCYV。 (C) 2013 Elsevier B.V. 保留所有权利。
A reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the detection of Cucurbit chlorotic yellows virus (CCYV). In this procedure, a set of four primers matching a total of six sequences in the coat protein gene region of CCYV was synthesized for the RT-LAMP assay using total RNA extracted from CCYV-infected melon leaf tissues, and the optimum reaction temperature and assay time were determined. The sensitivity assay showed that the virus was detectable in RT-LAMP reactions at dilutions of 1 x 10(-11), which was 10(5) times more sensitive than the RT-PCR assay. The RT-LAMP assay for CCYV and Sweet potato chlorotic stunt virus (SPCSV) exhibited high specificity for CCYV. This simple and sensitive method has potential for detection of CCYV in samples collected in the field. (C) 2013 Elsevier B.V. All rights reserved.