Infectious bronchitis virus detection in allantoic fluid using the polymerase chain reaction and a DNA probe.
A rapid extraction procedure was developed to purify infectious bronchitis virus (IBV) RNA from the allantoic fluid of inoculated embryonating eggs. Reverse transcription of viral RNA and the polymerase chain reaction (PCR) were used to amplify the viral genome from eight different strains of IBV comprising five different serotypes. A biotinylated DNA probe, prepared to a sequence within the PCR amplification product of the Beaudette strain of IBV, was used in a dot-hybridization assay; it detected the amplification products of all of the IBV strains examined. Reverse transcription and PCR amplification were judged to be specific for IBV. This was because amplification products were not detected by agarose gel electrophoresis or by dot-hybridization when template used in the PCR was extracted from allantoic fluid and the chorioallantoic membrane of uninoculated embryonating eggs or from allantoic fluid of embryonating eggs inoculated with other chicken upper respiratory viruses.
Jackwood
M W
MW
Department of Avian Medicine, College of Veterinary Medicine, University of Georgia, Athens 30605.
Kwon
H M
HM
Hilt
D A
DA
D000482:Allantois / Q000382:microbiology*; D000818:Animals; D001483:Base Sequence; D002642:Chick Embryo / Q000382:microbiology*; D015342:DNA Probes / Q000737:chemistry*; D004587:Electrophoresis, Agar Gel; D001351:Infectious bronchitis virus / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000737:chemistry* / Q000302:isolation & purification; D012680:Sensitivity and Specificity; D013698:Templates, Genetic
D015342:DNA Probes; D012367:RNA, Viral