Differentiation of Staphylococcus spp. by High-Resolution Melting Analysis
| Autoři | |
|---|---|
| Rok publikování | 2010 |
| Druh | Článek v odborném periodiku |
| Časopis / Zdroj | Canadian Journal of Microbiology |
| Fakulta / Pracoviště MU | |
| Citace | |
| Doi | http://dx.doi.org/10.1139/W10-091 |
| Obor | Genetika a molekulární biologie |
| Klíčová slova | HRMA; real-time PCR; 16S rRNA; bacterial detection |
| Popis | High-resolution melting analysis (HRMA) is a fast high-throughput method to scan for sequence variations in a target gene. The aim of this study was to test the potential of HRMA to distinguish particular bacterial species of the Staphylococcus genus even when using a broad-range PCR within the 16S rRNA gene where sequence differences are minimal. Genomic DNA samples isolated from 12 reference strains were subjected to a real-time PCR amplification of the 16S rRNA gene in the presence of fluorescent dye, followed by HRMA. Melting profiles were used as molecular fingerprints for bacterial species differentiation. HRMA of S. saprophyticus and S. xylosus resulted in undistinguishable profiles because of their identical sequences in the analyzed 16S rRNA region. The remaining reference strains were fully differentiated either directly or via high-resolution plots obtained by heteroduplex formation between coamplified PCR products of the tested staphylococcal strain and phylogenetically unrelated strain. |