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Viruses 2013, 5(9), 2272-2281; doi:10.3390/v5092272
Article

Development of a One-Step SYBR Green I Real-Time RT-PCR Assay for the Detection and Quantitation of Araraquara and Rio Mamore Hantavirus

†,* ,
,
,
 and
Virology Research Center, School of Medicine in Ribeirão Preto, University of São Paulo, Av. Bandeirantes n.3900, Monte Alegre, Ribeirão Preto, São Paulo14049-900, Brazil These authors contributed equally to this work.
* Author to whom correspondence should be addressed.
Received: 13 July 2013 / Revised: 10 September 2013 / Accepted: 17 September 2013 / Published: 19 September 2013
(This article belongs to the Special Issue Hantaviruses)
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Abstract

Hantaviruses are members of the family Bunyaviridaeandare an emergingcause of diseaseworldwidewithhighlethalityin the Americas. In Brazil, thediagnosis forhantaviruses is basedonimmunologictechniquesassociatedwithconventionalRT-PCR.Anovelone-stepSYBRGreenreal-timeRT-PCRwasdevelopedfor the detectionandquantitationofAraraquarahantavirus(ARAV)and Rio Mamore hantavirus (RIOMV). Thedetectionlimitof assay was 10copies/µLofRNA in vitro transcribed of segment S. The specificity of assay was evaluatedbymeltingcurveanalysis, which showed thattheAraraquaravirusamplifiedproductgenerate dameltpeak at80.83±0.89°C without generating primer-dimersornon-specificproducts.The assay was more sensitive than conventional RT-PCR and we detected two samples undetected by conventional RT-PCR. The one-step SYBR Green real-time quantitative RT-PCR is specific, sensible and reproducible, which makes it a powerful tool in both diagnostic applications and general research of ARAVand RIOMVand possibly other Brazilian hantaviruses.
Keywords: Hantaviruses; Araraquara virus; Rio Mamore virus; real-time quantitative RT-PCR; SYBR green I Hantaviruses; Araraquara virus; Rio Mamore virus; real-time quantitative RT-PCR; SYBR green I
This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).
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Machado, A.M.; de Souza, W.M.; de Pádua, M.; da Silva Rodrigues Machado, A.R.; Figueiredo, L.T.M. Development of a One-Step SYBR Green I Real-Time RT-PCR Assay for the Detection and Quantitation of Araraquara and Rio Mamore Hantavirus. Viruses 2013, 5, 2272-2281.

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