AN HOMOGENEOUS FLUORESCENCE POLYMERASE CHAIN-REACTION ASSAY TO IDENTIFY SALMONELLA

We have developed a semiquantitative homogeneous fluorescence assay th at combines polymerase chain reaction (PCR) amplification with direct fluorescence detection (HF-PCB). The assay eliminates the need to perf orm gel electrophoresis on test samples. Using a set of Salmonella-spe cific primers, this system was used to verify suspect colonies from cu lture plates as Salmonella. The fluorescence signal is generated by a nucleic acid dye, YO-PRO-1, that is included in the amplification reac tion. This homogeneous PCR assay was used to test 84 Salmonella strain s picked from selective culture plates. All data indicated positive re sults when compared with 17 non-Salmonella strains (in general, Citrob acter, Hafnia, Proteus, and Escherichia). The HF-PCR assay is a sensit ive, simple, accurate, and reproducible method that correlates well wi th size-exclusion high-performance liquid chromatography and gel elect rophoresis techniques as a means to monitor PCR-mediated DNA amplifica tion. This assay can confirm suspect colonies within 2.5 h. (C) 1997 A cademic Press.