Single-tube, non-interrupted reverse transcription PCR for detection of infectious pancreatic necrosis virus

Abstract

An assay using a single-tube, non-interrupted reverse transcription-polymerase chain reaction (RT-PCR) was established for the detection of infectious pancreatic necrosis virus (IPNV). Two primer sets, Pr D and Pr F, which framed a region within the gene coding for IPNV VP2 protein were used to amplify specific fragments of the IPNV genome. Amplified products were detected in cultured chinook salmon embryo cells (CHSE-2141 infected with different IPNV strain isolates including WB, Ja, Sp, Ab, VR299, 3372, MFK, and CV-HB-I, but not in uninfected CHSE-214 nor in cells infected with IHNV or infectious bursal disease virus {IBDV). The detection limit of this method was estimated with purified viral RNA from the WB strain of IPNV to be between 15 fg and 15 pg in ethidium bromide-stained gels.