Method of detection of classical swine fever

Details Method of detection of classical swine fever Method of detection of classical swine fever

C - Chemistry, Metallurgy

12

Q

C12Q 1/70 (2006.01) C07H 21/04 (2006.01) C07J 17/00 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 2597383

The invention relates to a multiplex real-time RT-PCR assay with a heterologous internal control system (i.e., EGFP-RNA) for the simple and fast diagnosis of classical swine fever virus (CSFV). Primers and FAM-labeled TaqMan probes, specific for CSFV were selected by analyzing the consensus sequence of the 5 '-non translated region of various CSFV strains. For determining the analytical sensitivity an in vitro transcript (T7-PC3Alf) of the 5' NTR was constructed and tested. Furthermore, a primer - probe system for the detection of the internal control sequence was established, and a multiplex assay using CSF-System 1 and the IC real-time PCR could be performed as a one-tube assay without loss of sensitivity or specificity.

L'invention concerne une bioanalyse de PCR quantitative en temps réel (RT-PCR) multiplex avec un système de régulation interne hétérologue (c.-à-d., EGFP-ARN) pour le diagnostic simple et rapide du virus de la fièvre porcine classique (CSFV). Des amorces et des sondes TaqMan à étiquette FAM, spécifiques du CSFV ont été sélectionnées par analyse de la séquence consensus de la région 5' non transduite de différentes souches du CSFV. Pour déterminer la sensibilité analytique, un transcrit in vitro (T7-PC3Alf) de 5' NTR a été construit et testé. En outre, un système d'amorce de sonde pour détecter la séquence de commande interne a été élaboré et une bioanalyse multiplex utilisant un système CSF 1 et la PCR en temps réel IC pourrait être effectuée sous forme de bioanalyse en un seul tube analyseur sans perte de sensibilité ou de spécificité.

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