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Diagn Microbiol Infect Dis. 2010 Jan;66(1):41-9. doi: 10.1016/j.diagmicrobio.2008.07.011. Epub 2008 Aug 21.

Activity and DNA contamination of commercial polymerase chain reaction reagents for the universal 16S rDNA real-time polymerase chain reaction detection of bacterial pathogens in blood.

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1
Institut für Angewandte Mikrobiologie und Biotechnologie, Hochschule Bremerhaven, An der Karlstadt 8, D-27568 Bremerhaven, Germany.

Abstract

Universal 16S rRNA gene polymerase chain reaction (PCR) is a promising means of detecting bacteremia. Among other factors, the PCR reagents play a prominent role for obtaining a high sensitivity of detection. The reagents are ideally optimized with respect to the amplifying activity and absence of contaminating DNA. In this study, it was shown in a universal 16S rDNA real-time PCR assay that commercial PCR reagents can vary greatly among each other in these characters. Only 1 of the 5 reagents tested met the criteria of sensitive detection of pathogen DNA with a minimum of false-positive results. The reagent was validated by the detection of pathogens at low titers using bacterial DNA extracted from blood that was spiked with various Gram-positive and Gram-negative bacteria.

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