Abstract：
:Detection of classical swine fever virus (CSFV) and its discrimination from other pestiviruses can be achieved by virus isolation (VI) in cell cultures, antigen detection, or molecular analysis. To simplify the latter, a 5'-nuclease assay (TaqMan) was developed for the rapid and specific detection of CSFV with the minimum of downstream PCR processing. A pair of 5'-non-coding region, panpestivirus-specific PCR primers were assessed in a one-step reverse transcription-PCR with each of 36 diverse pestiviruses. The PCR products were subsequently reamplified, in conjunction with a CSFV-specific fluorogenic probe, in a nested-PCR with a second set of panpestivirus PCR primers. During nested PCR, when the target of interest was present, the CSFV probe annealed to the amplicon between the forward and reverse primers and was subsequently cleaved via the 5'-3' nucleolytic activity of the DNA polymerase resulting in the release of the fluorescent reporter dye. Each PCR tube was then placed directly into a luminescence spectrometer to monitor for any increase in fluorescence due to cleavage of the probe. This assay detected representatives of all genetic sub-groups of CSFV, but gave negative results for other pestiviruses. A preliminary assessment showed that the method could be used to detect CSFV RNA extracted from infected pig blood with a sensitivity greater than that of VI.
journal_name
J Virol Methodsjournal_title
Journal of virological methodsauthors
McGoldrick A,Lowings JP,Ibata G,Sands JJ,Belak S,Paton DJdoi
10.1016/s0166-0934(97)00208-5subject
Has Abstractpub_date
1998-06-01 00:00:00pages
125-35issue
2eissn
0166-0934issn
1879-0984pii
S0166093497002085journal_volume
72pub_type
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