Abbas F, Andreasen J R, Jackwood M W
College of Veterinary Medicine, Oregon State University, Corvallis 97331, USA.
Avian Dis. 1996 Jan-Mar;40(1):56-62.
The polymerase chain reaction (PCR) was developed using infectious laryngotracheitis virus (ILTV) primers made from a portion of the ILTV thymidine kinase gene. DNA from various ILTV field isolates, from the USDA challenge strain of ILTV, and from commercial ILTV vaccines was specifically amplified. No amplification occurred using template DNA from uninfected chicken-embryo liver cells (CELC), several nonavian alphaher-pesviruses, Mycoplasma gallisepticum, Mycoplasma synoviae, Pasteurella hemolytica, Escherichia coli, a group I avian adenovirus, fowl poxvirus, or a psittacid herpesvirus. The 647-base pair-amplified ILTV PCR product was labeled to create a nonradioactive, biotinylated DNA probe. Hybridization using the probe detected ILTV DNA. Both PCR and hybridization yielded positive results with ILTV DNA but not with the DNA of other pathogens. Hybridization was specific for ILTV using a stringent salt solution for a 30-min wash step or a somewhat less stringent salt solution for a 60-min wash step. However, slight hybridization occurred with CELC DNA when the less stringent salt solution was used in a 30-min wash step.
聚合酶链反应（PCR）是利用从传染性喉气管炎病毒（ILTV）胸苷激酶基因的一部分制备的引物开发的。来自各种ILTV野外分离株、美国农业部ILTV攻毒株以及商业ILTV疫苗的DNA被特异性扩增。使用来自未感染鸡胚肝细胞（CELC）、几种非禽α疱疹病毒、鸡毒支原体、滑液支原体、溶血巴斯德菌、大肠杆菌、I群禽腺病毒、禽痘病毒或鹦鹉疱疹病毒的模板DNA未发生扩增。扩增的647碱基对的ILTV PCR产物被标记以制备非放射性生物素化DNA探针。使用该探针进行杂交可检测到ILTV DNA。PCR和杂交对ILTV DNA均产生阳性结果，但对其他病原体的DNA未产生阳性结果。使用严格盐溶液进行30分钟洗涤步骤或使用不太严格盐溶液进行60分钟洗涤步骤时，杂交对ILTV具有特异性。然而，当在30分钟洗涤步骤中使用不太严格盐溶液时，CELC DNA会发生轻微杂交。
