Polymerase chain reaction (PCR) detection of Listeria monocytogenes in diluted milk and reversal of PCR inhibition caused by calcium ions

• Published in: Letters in applied microbiology 1996-02, Vol.22 (2), p.153-158
• Main Authors: Bickley, J., Short, J.K., McDowell, D.G., Parkes, H.C.
• Format: Article
• Language: English
• Subjects: Animals; Base Sequence; Biological and medical sciences; Calcium; Cattle; Chelating Agents; DNA Primers - genetics; DNA, Bacterial - genetics; DNA, Bacterial - isolation & purification; Egtazic Acid; Food industries; Food Microbiology; Fundamental and applied biological sciences. Psychology; Humans; Listeria monocytogenes; Listeria monocytogenes - genetics; Listeria monocytogenes - isolation & purification; Listeriosis - etiology; Magnesium; Milk - chemistry; Milk - microbiology; Milk and cheese industries. Ice creams; Molecular Sequence Data; Polymerase Chain Reaction - methods; Polystyrenes; Polyvinyls
• ISSN: 0266-8254; 1472-765X
• DOI: 10.1111/j.1472-765X.1996.tb01131.x

J. BICKLEY, J. K. SHORT, D. G. MCDOWELL AND H. C. PARKES. 1996. DNA from Listeria monocytogenes was used as the model system from this investigation, with PCR primers based on the listeriolysin O gene. Under standard polymerase chain reaction (PCR) conditions and with no prior treatment, amplification failed in the presence of more than 5% milk. Since inhibition of the PCR occurred at the same milk concentrations with full fat, half fat and fat‐free milk, inhibition was not attributed to the fat content of the milk. Calcium ions were, however, identified as a major source of PCR inhibition. The results demonstrated that the inhibitory effects of calcium ions and milk could be partially reversed by increasing the magnesium concentration in the reaction to well above the standard levels normally required for PCR. This work has important implications for the use of the PCR in the direct detection of food pathogens.