Publication details
Vývoj a aplikace MOL-PCR pro detekci původců bakteriálních infekcí z potravin
| Title in English | Development and application of MOL-PCR for the detection of bacterial food-borne infections |
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| Authors | |
| Year of publication | 2018 |
| Type | Conference abstract |
| MU Faculty or unit | |
| Citation | |
| Description | Multiplex oligonucleotide ligation PCR assay (MOL-PCR) is one method that can be used to detect microbial pathogens. It allows analysis of multiple types of markers, such as unique sequences, insertions, deletions, or single nucleotide polymorphisms (SNPs) in one multiplex reaction. For detecting a specific target sequence, a pair of MOLigo probes is proposed. Each pair of MOLigo probes is specific to a particular target sequence, but all the pairs contain the same sequence for placement of universal primers. MOLigo probes are connected to the target sequence and ligated during a separate ligation step to form a complex ssDNA that serves as a template for subsequent amplification with universal pair of primers (one fluorescently labeled) during PCR. One of the MOLigo probes also contains so-called TAG,it is a unique sequence by which the PCR products hybridize to a bead with a covalently coupled anti-TAG sequence. Each set of beads therefore carries a different anti-TAG sequence complementary to the TAG sequence in the MOLigo probe, while each set of beads has a distinct spectral address given by the combination of red and infrared fluorophores by which the polystyrene beads are filled. The whole reaction and subsequent analysis of the individual sets of beads with the PCR products is carried out in a MAGPix instrument connected to a computer. MOL-PCR has been optimized for the detection of bacterial pathogens caused alimentary infections as Listeria monocytogenes, Escherichia coli, Yersinia enterocolitica, Salmonella enterica subs. enterica and Staphylococcus aureus. The versatility of MOL-PCR was used in their simultaneous detection. This work was created within the project of Security Research of the Ministry of the Interior of the Czech Republic VI 20152020044. |