Santurde G, Da Silva N, Villares R, Tabares E, Solana A, Bautista J M, Castro J M
Departamento de Patologia Animal L Universidad Complutense de Madrid. Facultad de Veterinaria, Madrid, Spain.
Vet Microbiol. 1996 Mar;49(1-2):81-92. doi: 10.1016/0378-1135(95)00169-7.
A procedure for direct detection of the BHV-1 genome in clinical samples, including semen, was developed. The method is based on the PCR amplification of a highly conserved DNA fragment within the glycoprotein gI sequence of the virus (323 bp between nt 1491 to nt 1814). The method is rapid and highly specific for all 27 subtypes assayed, which are included in the clinical and genetically different groups of BHV-1. The viral origin of the PCR product was assessed by Southern hybridization, with an internal probe. The method for DNA isolation from clinical samples included a fast extraction procedure with Chelex 100 resin allowing the loading of larger amounts of DNA in the PCR and in turn increasing the sensitivity of the method of detection. The level of sensitivity achieved by PCR was in the range of 1 TCID(50). This PCR assay may be an useful tool for BHV-1 monitoring in semen banks at low cost.
开发了一种直接检测临床样本（包括精液）中牛疱疹病毒1型（BHV - 1）基因组的方法。该方法基于对病毒糖蛋白gI序列内高度保守的DNA片段（核苷酸1491至1814之间的323 bp）进行PCR扩增。该方法快速且对所检测的所有27个亚型具有高度特异性，这些亚型包含在临床和基因不同的BHV - 1组中。通过用内部探针进行Southern杂交评估PCR产物的病毒来源。从临床样本中分离DNA的方法包括使用Chelex 100树脂的快速提取程序，该程序允许在PCR中加载更多量的DNA，进而提高检测方法的灵敏度。PCR达到的灵敏度水平在1个半数组织培养感染剂量（TCID50）范围内。这种PCR检测方法可能是以低成本对精液库中的BHV - 1进行监测的有用工具。
