Quantitative competitive reverse Transcription-PCR for quantification of dengue virus RNA

A quantitative competitive reverse transcription-PCR assay was developed to quantify dengue virus RNA in this study. The main features include a prime r pair targeting a highly conserved region in the capsid and the addition o f competing RNA that contains an internal deletion to provide a stringent i nternal control for quantification. It can be utilized to quantify RNA isol ated from the four dengue virus serotypes but not RNA isolated from other f laviviruses, including Japanese encephalitis virus and hepatitis C virus, b oth prevalent in Asia. It can also be used to quantify dengue virus RNA iso lated from the plasma of infected individuals. The sensitivity of the assay was estimated to be 10 to 50 copies of RNA per reaction, and twofold diffe rences in virus titer are distinguishable. This assay is a convenient, sens itive, and accurate method for quantification and can be used to further un derstanding of the pathogenesis of dengue virus infection.