Rapid purification of Salmonella DNA in minced meat and detection by real-time PCR

Four rapid and simple DNA purification and sample treatment protocols were evaluated for detection of Salmonella enterica in spiked minced meat, using a fluorogenic 5' nuclease (TaqMan) PCR assay in an ABI-Prism 7700 Sequence Detector. The detection limit with the single separation treatment of DNea sy was found to be 6-8 CFU in just 19 end-point fluorescence (C-t) values, while this was 22 C-t for a combination of DNeasy and BactXtractor. Extract ion by DNeasy resulted in C-t < 22, indicating a detection limit of < 10 Sa lmonella cells per 25 g, when the samples were inoculated with Salmonella b efore the overnight preenrichment. The method is currently being adapted to a BioRobot 3000 platform. However, the use of paramagnetic beads (DNA Dire ct) resulted in poor and variable detection limit.