Tham K M, Ng K, Young L W
Virology Section, Central Animal Health Laboratory, MAF Quality Management, New Zealand.
Arch Virol. 1994;135(3-4):355-64. doi: 10.1007/BF01310020.
A polymerase chain reaction (PCR) assay was developed for the detection of alcelaphine herpesvirus 1 (AHV1), a causative agent of malignant catarrhal fever (MCF) of ruminants. A pair of 20-base primers was constructed based on the published nucleotide sequence of gene A of the WC11 isolate of AHV1 and was used to amplify a DNA fragment of 413 base pairs. The optimised PCR assay was highly sensitive, i.e. it detected 10 fg of genomic DNA of AHV1 (WC11 isolate). The amplified fragment was shown to be specific for AHV1 DNA by (i) cleavage with XbaI which yielded 2 subfragments of approximately 140 and 280 base pairs and (ii) chemiluminescence Southern blot hybridisation with a digoxigenin-labelled 25-base internal probe. The PCR assay also amplified AHV1 gene sequences in tissue samples from deer and rabbits experimentally infected with materials derived from deer with clinical sheep-associated MCF.
已开发出一种聚合酶链反应（PCR）检测方法，用于检测反刍动物恶性卡他热（MCF）的病原体——非洲马瘟病毒1型（AHV1）。根据已发表的AHV1 WC11分离株基因A的核苷酸序列构建了一对20个碱基的引物，用于扩增一个413个碱基对的DNA片段。优化后的PCR检测方法具有高度敏感性，即它能检测到10 fg的AHV1（WC11分离株）基因组DNA。通过以下方式证明扩增片段对AHV1 DNA具有特异性：（i）用XbaI切割，产生约140和280个碱基对的2个亚片段；（ii）与地高辛标记的25个碱基内部探针进行化学发光Southern印迹杂交。该PCR检测方法还能扩增来自用与临床绵羊相关的MCF的鹿源材料实验感染的鹿和兔的组织样本中的AHV1基因序列。
