mericon DNA Bacteria Kit (100)

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Publication protocol

Detection S. typhimurium with real time PCR: Genomic DNA from captured S. typhimurium were directly extracted from tube. Three extraction techniques were employed following manufacturer instructions: heating extraction using mericon DNA bacteria kit (Qiagen) was used in Condition E, a magnetic extraction method using Dynabeads DNA direct universal kit (Thermo Fisher Scientific) was used in Condition D and ground chicken tests. DNA extraction was also done by heating at 100 °C in DI water for 10 minutes. Real time PCR was performed using the mericon Salmonella spp. kit (Qiagen). 10 µL of extracted DNA sample was mixed with 10 µL of mastermix (reconstituted mericon assay). A positive control with 10 µL purified salmonella DNA (provided with the kit) and a negative control with 10 µL dilution buffer (no DNA) were prepared as well. The PCR reactions were performed using a Rotor-gene-Q (Qiagen) RT PCR instrument. The sample was heated to 95 °C for 5 min. Then, up to 70 cycles were performed (each cycle included denaturation at 95 °C for 15 sec, annealing at 60 °C for 15 sec, and extension at 72 °C for 10 sec). The amplification was determined by monitoring the fluorescence of the FAM (495/520 nm excitation/emission) probe for the target gene and of the MAX (524/557 nm excitation/emission) probe for internal control

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Manufacturer protocol

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