EvaGreen-based real-time PCR assay for sensitive detection of enzootic nasal tumor virus 2

Mol Cell Probes. 2019 Apr:44:51-56. doi: 10.1016/j.mcp.2019.02.003. Epub 2019 Feb 13.

Abstract

Enzootic nasal tumor virus 2 (ENTV-2), the aetiological agent of enzootic nasal adenocarcinoma in goats, is prevalent in China; resulting in substantial economic losses to the goat-breeding industry. Therefore, it is necessary to establish an efficient detection method for the diagnosis and prevention of ENTV-2 infection. More recently, EvaGreen is emerging as a novel alternative fluorescent dye for quantitative real-time PCR because of its low cost, specific amplification and high resolution. In this study, we developed a specific, sensitive, and cost-effective detection method-an EvaGreen-based real-time PCR assay for the detection of ENTV-2. This assay exhibited high specificity and sensitivity and was able to detect ENTV-2 at concentrations as low as 3.0 × 101 copies, which was more sensitive than the conventional PCR method (detection limit, 3.0 × 102 copies). In addition, the reproducibility test indicated that EvaGreen dye in our assay had a good reproducibility. In conclusion, we report that a highly sensitive, specific, and cost-effective EvaGreen-based real-time PCR assay is successful for the rapid detection of ENTV-2.

Keywords: Enzootic nasal tumor virus 2; EvaGreen-based real-time PCR assay; Virus detection.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Betaretrovirus / genetics*
  • Betaretrovirus / isolation & purification*
  • DNA Primers / metabolism
  • Goats / virology
  • Limit of Detection
  • Nose Neoplasms / virology*
  • RNA, Viral / genetics
  • Real-Time Polymerase Chain Reaction / methods*
  • Reference Standards
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Zoonoses / virology*

Substances

  • DNA Primers
  • RNA, Viral