Development of a molecular-beacon-based multi-allelic real-time RT-PCR assay for the detection of human coronavirus causing severe acute respiratory syndrome (SARS-CoV): a general methodology for detecting rapidly mutating viruses
Development of a molecular-beacon-based multi-allelic real-time RT-PCR assay for the detection of human coronavirus causing severe acute respiratory syndrome (SARS-CoV): a general methodology for detecting rapidly mutating viruses
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DOI：
10.1007/s00705-010-0906-7
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发表时间：
2011-04-01
影响因子：
2.7
通讯作者：
Kostrikis, Leondios G.
中科院分区：
文献类型：
--
作者：
Hadjinicolaou, Andreas V.;Farcas, Gabriella A.;Kostrikis, Leondios G.
新出现的传染病已经引起了全球努力开发快速和准确的检测技术。病毒的快速突变性质是一个主要困难，突出了对遗传多样性菌株进行特异性检测的必要性。其中一种传染源是2003年出现的SARS相关冠状病毒（SARS-CoV）。本研究的目的是建立一种特异性的实时RT-PCR检测SARS-CoV，考虑到其固有的多态性的性质，由于遗传漂移和重组和连续和多次引入的可能性，遗传上不相同的菌株到人群中，通过使用错配容忍的分子信标设计，以特异性地检测SARS-CoV的S，E，M和N基因。这些应用于简单的，可重复的双重和多重实时PCR检测25个死后样本和构建的RNA控制，他们表现出较高的目标检测能力和特异性。这种检测方法可以很容易地适用于检测其他新出现的和快速突变的病原体。
Emerging infectious diseases have caused a global effort for development of fast and accurate detection techniques. The rapidly mutating nature of viruses presents a major difficulty, highlighting the need for specific detection of genetically diverse strains. One such infectious agent is SARS-associated coronavirus (SARS-CoV), which emerged in 2003. This study aimed to develop a real-time RT-PCR detection assay specific for SARS-CoV, taking into account its intrinsic polymorphic nature due to genetic drift and recombination and the possibility of continuous and multiple introductions of genetically non-identical strains into the human population, by using mismatch-tolerant molecular beacons designed to specifically detect the SARS-CoV S, E, M and N genes. These were applied in simple, reproducible duplex and multiplex real-time PCR assays on 25 post-mortem samples and constructed RNA controls, and they demonstrated high target detection ability and specificity. This assay can readily be adapted for detection of other emerging and rapidly mutating pathogens.