Uracil-DNA glycosylase-treated reverse transcription loop-mediated isothermal amplification for rapid detection of avian influenza virus preventing carry-over contamination

J Vet Sci. 2016 Sep 30;17(3):421-5. doi: 10.4142/jvs.2016.17.3.421.

Abstract

Here, we describe a uracil-DNA glycosylase (UNG)-treated reverse transcription loop-mediated isothermal amplification (uRT-LAMP) for the visual detection of all subtypes of avian influenza A virus (AIV). The uRT-LAMP assay can prevent unwanted amplification by carryover contamination of the previously amplified DNA, although the detection limit of the uRT-LAMP assay is 10-fold lower than that of the RT-LAMP without a UNG treatment. To the best of our knowledge, this is the first successful application of deoxyuridine triphosphate/UNG strategy in RT-LAMP for AIV detection, and the assay can be applied for the rapid, and reliable diagnosis of AIVs, even in contaminated samples.

Keywords: avian influenza virus; contamination; loop-mediated isothermal amplification; uracil-DNA glycosylase.

MeSH terms

  • Animals
  • Birds
  • DNA Contamination
  • Influenza A virus / isolation & purification*
  • Influenza in Birds / diagnosis*
  • Influenza in Birds / virology
  • Nucleic Acid Amplification Techniques / methods
  • Nucleic Acid Amplification Techniques / veterinary*
  • Reverse Transcription
  • Sensitivity and Specificity
  • Uracil-DNA Glycosidase / chemistry*

Substances

  • Uracil-DNA Glycosidase