Novel pan-serotype control RNA for dengue virus typing through real-time reverse transcription-polymerase chain reaction

J Virol Methods. 2019 Sep:271:113677. doi: 10.1016/j.jviromet.2019.113677. Epub 2019 Jun 10.

Abstract

Dengue virus (DENV) is the causative agent of one of the most important febrile illnesses worldwide. Four DENV serotypes are responsible for a broad clinical spectrum of the disease. Positive controls are costly and required for the validation of molecular test results of DENV serotyping. In this study, we describe the in silico design of the qDENV-Control plasmid with the target sequences to oligonucleotides and probes widely used for DENV serotyping, and the subsequent production of qDENV Control RNA by T7-driven run-off in vitro transcription. The qDENV Control RNA was successfully used to validate the positive and negative DENV serotyping results, allowing its incorporation in routine in-house protocols for virologic surveillance. This Control RNA allowed the absolute quantification of viral RNA copies from unknown samples as required in several fundamental studies.

Keywords: Dengue virus; In vitro transcription; Serotyping; qRT-PCR.

Publication types

  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Computer Simulation
  • DNA Primers / genetics
  • DNA Probes / genetics
  • Dengue / virology
  • Dengue Virus / classification*
  • Humans
  • RNA, Viral / analysis*
  • RNA, Viral / genetics*
  • Real-Time Polymerase Chain Reaction
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sensitivity and Specificity
  • Serogroup
  • Serotyping
  • Transcription, Genetic

Substances

  • DNA Primers
  • DNA Probes
  • RNA, Viral