Detection of bovine viral diarrhea (BVD) virus using the polymerase chain reaction.
The approach of reverse transcription (RT) followed by the polymerase chain reaction (PCR) was used to amplify three different fragments of the bovine viral diarrhea virus (BVDV) genome. Two sets of primers framed two different regions within the gene coding for protein p80, the third set of primers was selected to amplify cDNA within the envelope glycoprotein (gp53) region. All three sequences could be detected in the homologous strain (NADL), whereas only some of the fragments could be amplified in heterologous strains of BVDV. RNA extracted from infected cells as well as RNA extracted from viral particles could be detected using RT-PCR. The detection limit was 10(-1)-10(-2) TCID50 in ethidium bromide stained gels and could be further enhanced to 10(-2)-10(-4) TCID50 by hybridization after Southern transfer. The speed and the sensitivity of this method might be of relevance for diagnostic purposes as well as for studies on epidemiology and pathogenesis of infection with BVD virus.
Hertig
C
C
Institute of Veterinary Virology, University of Bern, Switzerland.
Pauli
U
U
Zanoni
R
R
Peterhans
E
E
D000818:Animals; D001483:Base Sequence; D015139:Blotting, Southern; D002417:Cattle; D015342:DNA Probes; D004279:DNA, Viral / Q000737:chemistry*; D001908:Diarrhea Viruses, Bovine Viral / Q000235:genetics / Q000302:isolation & purification*; D005784:Gene Amplification; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D011237:Predictive Value of Tests; D012367:RNA, Viral / Q000032:analysis*; D015183:Restriction Mapping; D014158:Transcription, Genetic
D015342:DNA Probes; D004279:DNA, Viral; D012367:RNA, Viral