Development of a primer-probe energy transfer based real-time PCR for detection of Marek's disease virus

Annette Malene Barfoed, E. Østergaard, P.L. Frandsen, E.B. Nielsen, E. Sandberg, Thomas Bruun Rasmussen

    Research output: Contribution to journalJournal articleResearchpeer-review

    Abstract

    A real-time PCR assay, which enables simultaneous detection and differentiation of all three serotypes of Marek's disease virus, without the need for post-PCR sequencing, has been developed. The assay is based on the primer-probe energy transfer real-time PCR, which has a relatively high tolerance towards point mutations in the probe region. The PCR is followed by a probe melting point analysis, which enables confirmation of identity of amplicon and differentiation of serotypes. The assay targets the MDV031 gene, encoding UL19 major capsid protein-like protein and was shown to be quantitative, with a detection limit below 10 TCID50/ml starting material. This sensitivity is similar to the one obtained with traditional virus cultivation. However, the PCR method can provide a laboratory result within a day, while the virus cultivation method takes more than a week to perform. The new method will be useful for testing of avian live viral vaccines and screening for extraneous agents.
    Original languageEnglish
    JournalJournal of Virological Methods
    Volume165
    Issue number1
    Pages (from-to)21-26
    ISSN0166-0934
    DOIs
    Publication statusPublished - 2010

    Keywords

    • Real-time PCR
    • Marek's disease virus
    • Primer-probe energy transfer

    Cite this