Detection of strawberry RNA and DNA viruses by RT-PCR using total nucleic acid as a template
Detection of strawberry RNA and DNA viruses by RT-PCR using total nucleic acid as a template
复制标题
DOI：
10.1111/j.1439-0434.2007.01254.x
复制
发表时间：
2007-08-01
影响因子：
1.5
通讯作者：
Dai, H.
中科院分区：
文献类型：
--
作者：
Chang, L.;Zhang, Z.;Dai, H.
草莓轻度黄边病毒(SMYEV)、草莓斑驳病毒(SMoV)和草莓脉带病毒(SVBV)这三种草莓病毒可造成全球商业草莓的严重损失。本文以草莓叶总核酸和总RNA为模板，通过单次和多重逆转录聚合酶链反应（RT-PCR）检测上述病毒进行比较。当上样到琼脂糖凝胶上时，使用改良 CTAB 方法提取的总核酸以及使用改良 RNeasy 方法和改良 CTAB 方法提取的总 RNA 提取物通过电泳进行检测，得到高质量的核糖体 28S 和 18S RNA。以总RNA和总核酸为模板时，无论是单重RT-PCR还是双重RT-PCR，SMYEV和SMoV检测结果相同，扩增条带强度基本相同。以总核酸为模板，多重RT-PCR检测SMYEV和SMoV的结果与单重和双重RT-PCR扩增的结果吻合较好，而SVBV则与以纯DNA为模板直接PCR扩增的检测结果相似。本研究表明，通过改进的 CTAB 方法从草莓叶中提取的总核酸适合作为 RT-PCR 病毒检测的模板。这使得草莓病毒的流行病学调查变得更加容易。
Strawberry mild yellow edge virus (SMYEV), Strawberry mottle virus (SMoV) and Strawberry vein banding virus (SVBV), three of strawberry viruses can cause a severe loss in commercial strawberry worldwide. In this paper, total nucleic acid from strawberry leaves was compared with total RNA as the template for detection of the above viruses both by single and multiplex reverse transcription polymerase chain reaction (RT-PCR). Total nucleic acid extract with the modified CTAB method and total RNA extracts with both the modified RNeasy method and the modified CTAB method examined by electrophoresis resulted in a high quality ribosomal 28S and 18S RNA when loaded on an agarose gel. When total RNA and total nucleic acid were used as templates, detection results of SMYEV and SMoV were identical and the amplified band intensity was nearly the same whether by single or duplex RT-PCR. Using total nucleic acid as the template, the detection results of SMYEV and SMoV by multiplex RT-PCR were in a good concord with that amplified by single and duplex RT-PCR, and for SVBV this was similar to the result of detection that amplified by direct PCR using pure DNA as the template. This study demonstrates that total nucleic acid extracted from strawberry leaves by a modified CTAB method is suitable as a template for virus detection by RT-PCR. This makes investigation of the epidemiology of strawberry viruses much easier.