Research on the quantification method to detect pathogenic bacteria in instant rice noodles by 3-plex droplet digital polymerase chain reaction test

ObjectiveThis study aimed to establish a quantitative 3-plex droplet digital polymerase chain reaction （ddPCR） method for simultaneously detecting the copy numbers of Salmonella， Bacillus cereus， and Listeria monocytogenes in instant food.MethodsThree pairs of primers and probes corresponding to three single-copy-genes were selected as target genes. The genes were the essC gene in Bacillus cereus， ttrA/ttrC gene in Salmonella， and invasion-associated endopeptidase gene in Listeria monocytogenes. The specificity of the primers and probes were verified by real-time fluorescence quantitative PCR separately. A 3-plex ddPCR method was constructed to detect the copy numbers of three pathogenic bacteria simultaneously.ResultsThe linear ranges were： 25-22 687 copies/20 µL for Salmonella， 19-15 620 copies/20 µL for Bacillus cereus， and 18-23 373 copies/20 µL for Listeria monocytogenes. The three linear correlation coefficients were r≥0.999. relative standard deviation （RSD）≤12% at six concentrations and repeated thric