An alternative method for the synthesis of competitor RNA transcripts useful for specific detection and quantitation of dengue virus serotype 2 genome and replicative intermediate RNA

Abstract

The development of a quantitative-competitive reverse transcription-PCR (RT-PCR) assay to quantify dengue virus (DEN) genome (vRNA) and its replicative intermediate RNA (vRI) is described. A highly conserved region located on the DEN capsid-premembrane genes was used to produce a competitor RNA molecule which contains an internal deletion of 70 nucleotides. The competitor provides a suitable internal control useful to quantify viral RNA from all four dengue virus (DEN 1-4) serotypes. The detection limit of the assay was found to be 100 copies per reaction. This is a rapid, simple, sensitive, inexpensive and easy method for quantitation of DEN RNA species.