Comparison of three rapid and easy bacterial DNA extraction methods for use with quantitative real-time PCR.

ABSTRACT: The development of fast and easy on-site molecular detection and quantification methods for hazardous microbes on solid surfaces is desirable for several applications where specialised laboratory facilities are absent. The quantification of bacterial contamination necessitates the assessment of the efficiency of the used methodology as a whole, including the preceding steps of sampling and sample processing. We used quantitative real-time polymerase chain reaction (qrtPCR) for Escherichia coli and Staphylococcus aureus to measure the recovery of DNA from defined numbers of bacterial cells that were subjected to three different DNA extraction methods: the QIAamp DNA Mini Kit, Reischl et al.'s method and FTA Elute. FTA Elute significantly showed the highest median DNA extraction efficiency of 76.9% for E. coli and 108.9% for S. aureus. The Reischl et al. method and QIAamp DNA Mini Kit inhibited the E. coli qrtPCR assay with a 10-fold decrease of detectable DNA. None of the methods inhibited the S. aureus qrtPCR assay. The FTA Elute applicability was demonstrated with swab samples taken from the International Space Station (ISS) interior. Overall, the FTA Elute method was found to be the most suitable to selected criteria in terms of rapidity, easiness of use, DNA extraction efficiency, toxicity, and transport and storage conditions.

SUBMITTER: van Tongeren SP 

PROVIDER: S-EPMC3181010 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature