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OJVRTM
Online Journal of Veterinary Research©
Volume 19(7): 471-479, 2015.
Most Probable Number PCR and real-time
PCR assays for detection of Listeria monocytogenes in milk
Elham Karimi a(DVM), Abdolah Jamshidi a*(DVM, PhD), Saeid
Khanzadi a(DVM, PhD), Mohammad Reza
Bassami b(DVM, PhD), Amin Nozari b(DVM
)
aFood Hygiene, bBiotechnology,
Faculty of Veterinary Medicine, Ferdowsi University of Mashhad, Mashhad, Iran.
ABSTRACT
Karimi E, JamshidiA,
Khanzadi S, Bassami MR, Nozari
A., Most Probable Number PCR and real-time PCR assays for detection of Listeria
monocytogenes in milk, OnlJ Vet Res., 19(7): 471-479,
2015. Results from most Probable Number (MPN)-PCR and real-time (RT)PCR assays without
pre-enrichment for detection of Listeria
monocytogenes in milk are presented. L. monocytogenes pure culture and 8
other background bacterial suspensions of 107 and 101
CFU/ml were prepared. Three test tube MPN sets were prepared from 101
CFU/ml of bacterial mix suspension in Listeria
enrichment broth. After incubation, turbid MPN tubes were subjected to DNA
isolation and MPN-PCR assay. For RT-PCR assay, 9 ten-fold dilutions were
prepared from genomic DNA isolate from bacterial mix suspension and L.
monocytogenes stock suspension (107-10-2 genomic DNA
copy/ml) as test and standard samples, respectively. MPN-PCR and RT-PCR assays
were conducted in triplicate using appropriate primers for the prfA gene. According to our findings the detection
limit of MPN-PCR was more precise than RT-PCR (101 CFU/ml and 104
CFU/ml, respectively). The finding suggests
that MPN-PCR could be a better alternative
than RT-PCR without any pre-enrichment, for analysis of low levels of L. monocytogenes in food samples.
Keywords: L. monocytogenes, MPN-PCR, real-time
PCR.
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