Sap-direct RT-PCR for the rapid detection of coleus blumei viroids of the genus Coleviroid from natural host plants
Sap-direct RT-PCR for the rapid detection of coleus blumei viroids of the genus Coleviroid from natural host plants
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DOI：
10.1016/j.jviromet.2011.03.018
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发表时间：
2011-06-01
影响因子：
3.1
通讯作者：
Li, Shifang
中科院分区：
文献类型：
--
作者：
Jiang, Dongmei;Wu, Zujian;Li, Shifang
建立了一种快速检测Coleviroid属3种类病毒的快速检测方法。利用分子生物学实验室常用的移液器直接从彩叶草汁液中获得cDNA合成模板，并根据彩叶草类病毒中央保守区（CCR）序列设计一对通用引物，同时检测3种彩叶草类病毒（CbVds）。RT-PCR结果表明，CbVd-1，CbVd-5，CbVd-6可以准确地检测到类病毒感染的植物，但不能在无类病毒的植物。RT-PCR、斑点杂交、测序和批量检测结果表明，该方法可用于快速鉴定CbVds。该方法还将不同样品之间的交叉污染降至最低。认为该技术是一种快速、简便的鉴定和克隆CbVds的有效方法。皇冠版权所有（C）2011由Elsevier B. V.出版。保留所有权利。
A simple and fast sap-direct RT-PCR (reverse transcription-polymerase chain reaction) for the rapid detection of 3 viroids of the genus Coleviroid is presented. The templates for cDNA synthesis were obtained directly from the sap of coleus using a pipettor, a common tool in molecular biology laboratories, and 3 coleus blumei viroids (CbVds) were detected simultaneously using a pair of universal primers designed according to sequences in the central conserved region (CCR) of CbVds. RT-PCR results demonstrated that CbVd-1, CbVd-5, and CbVd-6 can be detected accurately in viroid-infected plants but not in viroid-free plants. The results of RT-PCR, dot-blot, sequencing, and batch-detection revealed that this method can be used to identify CbVds rapidly. The method also reduces cross-contamination among different samples to a minimum. It is considered that this rapid and simple technique is an effective method for the identification and cloning of CbVds. Crown Copyright (C) 2011 Published by Elsevier B.V. All rights reserved.