PCR detection of nearly any dengue virus strain using a highly sensitive primer 'cocktail'

Charul Gijavanekar, Maria Añez-Lingerfelt, Chen Feng, Catherine Putonti, George E. Fox, Aniko Sabo, Yuriy Fofanov, Richard C. Willson

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

PCR detection of viral pathogens is extremely useful, but suffers from the challenge of detecting the many variant strains of a given virus that arise over time. Here, we report the computational derivation and initial experimental testing of a combination of 10 PCR primers to be used in a single high-sensitivity mixed PCR reaction for the detection of dengue virus. Primer sequences were computed such that their probability of mispriming with human DNA is extremely low. A 'cocktail' of 10 primers was shown experimentally to be able to detect cDNA clones representing the four serotypes and dengue virus RNA spiked into total human whole blood RNA. Computationally, the primers are predicted to detect 95% of the 1688 dengue strains analyzed (with perfect primer match). Allowing up to one mismatch and one insertion per primer, the primer set detects 99% of strains. Primer sets from three previous studies have been compared with the present set of primers and their relative sensitivity for dengue virus is discussed. These results provide the formulation and demonstration of a mixed primer PCR reagent that may enable the detection of nearly any dengue strain irrespective of serotype, in a single PCR reaction, and illustrate an approach to the broad problem of detecting highly mutable RNA viruses. PCR detection of dengue virus suffers from the challenge of detecting the many variant strains which arise over time. We report computational derivation and experimental testing of a combination of ten PCR primers used in a single PCR reaction, for detection of nearly any of the thousands of known dengue virus strains irrespective of serotype

Original languageEnglish (US)
Pages (from-to)1676-1687
Number of pages12
JournalFEBS Journal
Volume278
Issue number10
DOIs
StatePublished - May 2011
Externally publishedYes

Fingerprint

Dengue Virus
Viruses
Polymerase Chain Reaction
RNA
Dengue
Testing
Pathogens
RNA Viruses
Blood
Demonstrations
Complementary DNA
Clone Cells
DNA

Keywords

  • cocktail PCR
  • dengue virus
  • diagnostic
  • PCR
  • primer

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Gijavanekar, C., Añez-Lingerfelt, M., Feng, C., Putonti, C., Fox, G. E., Sabo, A., ... Willson, R. C. (2011). PCR detection of nearly any dengue virus strain using a highly sensitive primer 'cocktail'. FEBS Journal, 278(10), 1676-1687. https://doi.org/10.1111/j.1742-4658.2011.08091.x

PCR detection of nearly any dengue virus strain using a highly sensitive primer 'cocktail'. / Gijavanekar, Charul; Añez-Lingerfelt, Maria; Feng, Chen; Putonti, Catherine; Fox, George E.; Sabo, Aniko; Fofanov, Yuriy; Willson, Richard C.

In: FEBS Journal, Vol. 278, No. 10, 05.2011, p. 1676-1687.

Research output: Contribution to journalArticle

Gijavanekar, C, Añez-Lingerfelt, M, Feng, C, Putonti, C, Fox, GE, Sabo, A, Fofanov, Y & Willson, RC 2011, 'PCR detection of nearly any dengue virus strain using a highly sensitive primer 'cocktail'', FEBS Journal, vol. 278, no. 10, pp. 1676-1687. https://doi.org/10.1111/j.1742-4658.2011.08091.x
Gijavanekar C, Añez-Lingerfelt M, Feng C, Putonti C, Fox GE, Sabo A et al. PCR detection of nearly any dengue virus strain using a highly sensitive primer 'cocktail'. FEBS Journal. 2011 May;278(10):1676-1687. https://doi.org/10.1111/j.1742-4658.2011.08091.x
Gijavanekar, Charul ; Añez-Lingerfelt, Maria ; Feng, Chen ; Putonti, Catherine ; Fox, George E. ; Sabo, Aniko ; Fofanov, Yuriy ; Willson, Richard C. / PCR detection of nearly any dengue virus strain using a highly sensitive primer 'cocktail'. In: FEBS Journal. 2011 ; Vol. 278, No. 10. pp. 1676-1687.
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