A fast and robust method for full genome sequencing of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) Type 1 and Type 2

Lise Kirstine Kvisgaard, Charlotte Kristiane Hjulsager, Ulrik Fahnøe, Solvej Østergaard Breum, Tahar Ait-Ali, Lars Erik Larsen

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    Abstract

    PRRSV is a positive-sense RNA virus with a high degree of genetic variability among isolates. For diagnostic sensitivity and vaccine design it is essential to monitor PRRSV genetic diversity. However, to date only a few full genome sequences of PRRSV isolates have been made publicly available. In the present study, fast and robust methods for long range RT-PCR amplification and subsequent next generation sequencing (NGS) were developed and validated on nine Type 1 and nine Type 2 PRRSV viruses. The methods generated robust and reliable sequences both on primary material and cell culture adapted viruses and the protocols performed well on all three NGS platforms tested (Roche 454 FLX, Illumina HiSeq2000, and Ion Torrent PGM™ Sequencer). These methods will greatly facilitate the generation of more full genome PRRSV sequences globally.
    Original languageEnglish
    JournalJournal of Virological Methods
    Volume193
    Issue number2
    Pages (from-to)697-705
    ISSN0166-0934
    DOIs
    Publication statusPublished - 2013

    Keywords

    • PRRSV
    • Full genome sequencing
    • Next generation sequencing
    • Illumina HiSeq2000
    • Roche 454 FLX
    • Ion Torrent PGM sequencer

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