Comparison of dot blot hybridization, polymerase chain reaction, and virus isolation for detection of bovine herpesvirus-1 (BHV-1) in artificially infected bovine semen.

摘要: Bovine semen samples spiked with bovine herpesvirus 1 (BHV-1) were used to compare dot blot hybridization, polymerase chain reaction (PCR), and virus isolation for detection of BHV-1 in semen. The PCR amplification primers targeting the thymidine kinase gene a nucleic acid releasing cocktail (GeneReleaser); product was as DNA probe hybridization; done primary fetal testis (BFT) cell cultures. Semen diluted 1:20 tissue culture medium had least cytotoxicity inhibition viral cytopathic effects BFT cells, allowing TCID50/100 microL suspension by isolation. presence foreign such sperm or salmon increased sensitivity hybridization detecting BHV-1, 20,000 neat eliminated diluting medium. best obtained when buffer pH 9.0, 1.0 mM MgCl2 used. Under these conditions, followed ethidium bromide staining agarose gels could detect TCID20/100 sample, whereas Southern 0.01 sample.(ABSTRACT TRUNCATED AT 250 WORDS)