کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
8747015 | 1593418 | 2018 | 17 صفحه PDF | دانلود رایگان |
عنوان انگلیسی مقاله ISI
Development of a droplet digital RT-PCR for the quantification of foot-and-mouth virus RNA
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کلمات کلیدی
موضوعات مرتبط
علوم زیستی و بیوفناوری
ایمنی شناسی و میکروب شناسی
ویروس شناسی
پیش نمایش صفحه اول مقاله
چکیده انگلیسی
Foot-and-mouth-disease (FMD) is a highly contagious disease of domestic animals which can result in substantial economic losses, caused by the FMD virus (FMDV). The aim of this study was to develop and standardize a novel reverse transcriptase droplet digital PCR (RT-ddPCR) assay for the quantification of FMDV RNA. This assay was based upon an OIE-recognized real-time RT-PCR that detects the 3D-encoding region of FMDV. The limit of detection at 101.4 TCID50/mL and 26.5 copies was determined using FMDV-A24-Cruzeiro-virus and a plasmid containing the 3D-FMDV sequences, respectively. FMDV O, A and C serotypes and 11 species of non-FMDV were used to confirm the sensitivity and specificity of the assay. The RT-ddPCR was standardized using 60 bovine samples (representing negative and positive samples of epithelium and/or oesophageal-pharyngeal [OP] fluid) from animals suspected of vesicular diseases and previously tested by RT-qPCR. The RT-ddPCR showed robustness, sensitivity, specificity and accuracy, with similar results to the RT-qPCR. Moreover, the new RT-ddPCR diagnostic tool allowed the absolute quantification of FMDV RNA from epithelium and OP-fluid samples, as well as having the advantages of direct quantification by endpoint, eliminating the need for a calibration standard curve required in quantitative real-time RT-PCR.
ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Journal of Virological Methods - Volume 259, September 2018, Pages 129-134
Journal: Journal of Virological Methods - Volume 259, September 2018, Pages 129-134
نویسندگان
Tatiana F. Pinheiro-de-Oliveira, Antonio A. Jr, Marcelo F. Camargos, Mateus Laguardia-Nascimento, Anapolino M. de Oliveira, Ana C.P. Cottorello, Aristoteles Goes-Neto, Edel F. Barbosa-Stancioli,