基于发夹探针的环环扩增的用于 RNA 病毒检测的比色核酸检测分析。
Colorimetric nucleic acid testing assay for RNA virus detection based on circle-to-circle amplification of padlock probes.
机构信息
Department of Immunology, Genetics and Pathology, The Rudbeck Laboratory, Uppsala University, 75185 Uppsala, Sweden.
出版信息
J Clin Microbiol. 2011 Dec;49(12):4279-85. doi: 10.1128/JCM.00713-11. Epub 2011 Sep 28.
Abstract
We developed a molecular diagnostic method for detection of RNA virus based on padlock probes and colorimetric readout. The feasibility of our approach was demonstrated by using detection of Crimean-Congo hemorrhagic fever (CCHF) virus as a model. Compared with conventional PCR-based methods, our approach does not require advanced equipment, involves easier assay design, and has a sensitivity of 10(3) viral copies/ml. By using a cocktail of padlock probes, synthetic templates representing different viral strain variants could be detected. We analyzed 34 CCHF patient samples, and all patients were correctly diagnosed when the results were compared to those of the current real-time PCR method. This is the first time that highly specific padlock probes have been applied to detection of a highly variable target sequence typical of RNA viruses.
摘要
我们开发了一种基于发夹探针和比色读出的用于检测 RNA 病毒的分子诊断方法。我们的方法的可行性通过以克里米亚-刚果出血热（CCHF）病毒作为模型来证明。与基于常规 PCR 的方法相比，我们的方法不需要先进的设备，涉及到更简单的测定设计，并且具有 10^3 个病毒拷贝/ml 的灵敏度。通过使用发夹探针的混合物，可以检测代表不同病毒株变体的合成模板。我们分析了 34 个 CCHF 患者样本，当将结果与当前实时 PCR 方法进行比较时，所有患者的诊断结果均正确。这是首次将高度特异性的发夹探针应用于检测 RNA 病毒典型的高度变异靶序列。
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