Jackwood M W, Kwon H M, Hilt D A
Department of Avian Medicine, College of Veterinary Medicine, University of Georgia, Athens 30605.
Avian Dis. 1992 Apr-Jun;36(2):403-9.
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.
开发了一种快速提取程序，用于从接种的鸡胚尿囊液中纯化传染性支气管炎病毒（IBV）RNA。利用病毒RNA的逆转录和聚合酶链反应（PCR）从包括五种不同血清型的八种不同IBV毒株中扩增病毒基因组。制备了针对IBV Beaudette毒株PCR扩增产物内一个序列的生物素化DNA探针，用于斑点杂交试验；它检测到了所有检测的IBV毒株的扩增产物。逆转录和PCR扩增被判定对IBV具有特异性。这是因为当PCR中使用的模板从未接种的鸡胚尿囊液和绒毛尿囊膜或接种了其他鸡上呼吸道病毒的鸡胚尿囊液中提取时，琼脂糖凝胶电泳或斑点杂交均未检测到扩增产物。
