Raue R, Hess M
Institut für Geflügelkrankheiten, Freie Universität Berlin, Germany.
J Virol Methods. 1998 Aug;73(2):211-7. doi: 10.1016/s0166-0934(98)00065-2.
Three different polymerase chain reactions (PCRs), two of them combined with restriction enzyme analysis (REA), were developed for detection and differentiation of all 12 fowl adenovirus (FAV) serotypes and the egg drop syndrome (EDS) virus. For primer construction FAV1, FAV10 and EDS virus hexon proteins were aligned and conserved and variable regions were determined. Two primer sets (H1/H2 and H3/H4) for single use were constructed which hybridize in three conserved regions of hexon genes. Each primer pair amplifies approximately half of the hexon gene including two loop regions. An amplification product was detected with both primer sets using purified DNA from all FAV1-12 reference strains. Viral EDS DNA was negative using the H1/H2 or H3/H4 primer pair. HaeII digestion of the H1/H2 amplification products differentiates between all viruses except FAV4 and FAV5. In comparison, much more clustering among genomic closely related FAV serotypes was seen after HpaII digestion of the H3/H4 PCR products. Oligonucleotides H5/H6 located in the variable regions of EDS virus hexon gene do not detect any of the FAV serotypes. The PCRs and REA described are suitable to detect all avian adenoviruses infecting chickens, to distinguish all 12 FAV reference strains and to differentiate FAVs from the EDS virus.
开发了三种不同的聚合酶链反应（PCR），其中两种与限制性内切酶分析（REA）相结合，用于检测和区分所有12种禽腺病毒（FAV）血清型和减蛋综合征（EDS）病毒。为构建引物，对FAV1、FAV10和EDS病毒六邻体蛋白进行了比对，确定了保守区和可变区。构建了两个单独使用的引物组（H1/H2和H3/H4），它们在六邻体基因的三个保守区杂交。每个引物对扩增大约一半的六邻体基因，包括两个环区。使用来自所有FAV1 - 12参考毒株的纯化DNA，两个引物组均检测到扩增产物。使用H1/H2或H3/H4引物对，病毒EDS DNA呈阴性。H1/H2扩增产物的HaeII消化可区分除FAV4和FAV5之外的所有病毒。相比之下，H3/H4 PCR产物经HpaII消化后，在基因组密切相关的FAV血清型之间观察到更多聚类现象。位于EDS病毒六邻体基因可变区的寡核苷酸H5/H6未检测到任何FAV血清型。所描述的PCR和REA适用于检测感染鸡的所有禽腺病毒，区分所有12种FAV参考毒株，并区分FAV与EDS病毒。
