Wagter L H, Glas R D, Bleumink-Pluym N, Van Engelenburg F A, Rijsewijk F A, Houwers D J
Animal Health Service, Drachten, The Netherlands.
Vet Res Commun. 1996;20(4):401-8. doi: 10.1007/BF00366546.
A PCR assay for the detection of bovine herpesvirus type 1 (BHV1) DNA in selectively digested whole bovine semen was developed and evaluated. A brief treatment with proteinase-K was used to lyse free virus, virus present in non-sperm cells and virus adhering to the spermatozoa. Genomic bovine DNA was not released by this treatment. Primers and probes were based on the nucleotide sequence of the gD gene. BHV1 virus-spiked split samples were used as positive controls and the PCR products were detected by eye in ethidium bromide-stained agarose gels. Sequentially collected non-extended semen samples from experimentally infected bulls were used to compare this assay with virus isolation. Of a total of 162 ejaculates, 51 were found positive by virus isolation, whereas PCR detected BHV1 DNA in 73. PCR detected BHV1 DNA for a longer period after infection and reactivation. Apart from its superior sensitivity, this PCR assay also has the advantage of being a relatively simple procedure, providing results within 24 h.
开发并评估了一种用于检测选择性消化的全牛精液中牛疱疹病毒1型（BHV1）DNA的聚合酶链反应（PCR）检测方法。用蛋白酶K进行短暂处理，以裂解游离病毒、非精子细胞中存在的病毒以及附着在精子上的病毒。这种处理不会释放基因组牛DNA。引物和探针基于gD基因的核苷酸序列。用添加了BHV1病毒的分割样本作为阳性对照，并通过在溴化乙锭染色的琼脂糖凝胶中肉眼观察来检测PCR产物。使用从实验感染公牛中顺序收集的未稀释精液样本，将该检测方法与病毒分离法进行比较。在总共162份射精样本中，病毒分离法检测出51份呈阳性，而PCR检测出73份含有BHV1 DNA。PCR在感染和再激活后更长时间内检测到BHV1 DNA。除了具有更高的灵敏度外，这种PCR检测方法还具有操作相对简单的优点，可在24小时内得出结果。
