A more sensitive and rapid multiplex RT-PCR assay combining with magnetic nanobeads for simultaneous detection of viruses in sweet potato
A more sensitive and rapid multiplex RT-PCR assay combining with magnetic nanobeads for simultaneous detection of viruses in sweet potato
复制标题
更灵敏、更快速的多重 RT-PCR 检测结合磁纳米珠同时检测甘薯中的病毒
DOI：
10.1007/s10658-014-0447-0
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发表时间：
2014-09-01
影响因子：
1.8
通讯作者：
Lin, Hong-Hui
中科院分区：
文献类型：
--
作者：
Deng, Xing-Guang;Zhu, Feng;Lin, Hong-Hui
建立了一种改良的复合RT-PCR-磁性纳米微球（MNB-RT-PCR）方法，用于甘薯G病毒（SPVG）、甘薯羽状斑驳病毒（SPFMV）、甘薯C病毒（SPVC）和甘薯褪绿斑点病毒（SPCFV）的同时检测。设计了4对引物，分别扩增出SPVG、SPFMV、SPVC和SPCFV的169、357、516和900 bp的PCR产物。使用不同的病毒模板组合测试该方法的特异性，并通过测序确认扩增产物的身份。通过单一和多重MNB-RT-PCR检测所有四种病毒的检测限相当。与传统的CTAB-RT-PCR检测方法进行比较，结果表明MNB-RT-PCR检测方法更快速、灵敏。结果表明，多重MNB-RT-PCR技术是甘薯病毒检测的有效方法。
An improved multiplex RT-PCR assay combined with magnetic nanobeads (MNB-RT-PCR) was developed for simultaneous detection of four sweet potato viruses,Sweet potato virus G(SPVG),Sweet potato feathery mottle virus(SPFMV),Sweet potato virus C(SPVC) andSweet potato chlorotic fleck virus(SPCFV). Four primer pairs specific for each virus were designed and the corresponding PCR products were 169, 357, 516 and 900 bp in length for SPVG, SPFMV, SPVC and SPCFV, respectively. The specificity of the method was tested using different combinations of virus templates, and the identities of the amplification products were confirmed by sequencing. The limits of detection for all four viruses by single and multiplex MNB-RT-PCR assays were comparable. The assay was further evaluated using laboratory and field samples compared with a conventional CTAB-RT-PCR assay, and the comparative results showed that the MNB-RT-PCR assay was more rapid and sensitive. These results suggest that the multiplex MNB-RT-PCR assay is an effective and preferable method for virus detection in sweet potato.