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

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.