Introduction
Staphylococcus aureus is one of the most successful human pathogens, which can be detected in varied parts of the world . ()Some factors including virulence factor and drug resistance are efficacious in increasing infectious related death rate. Firstly, Methicillin-resistant Staphylococcus aureus (MRSA) strains were recognized in the early 1960s, moreover, they considered as a nosocomial deathful pathogen due to their role in increasing costs and bedridden length() In 2010 almost 15-20 % of patients with Staphylococcus aureus bacteremia were dead.() One of the most important ways in controlling and preventing of hospital infections is discerning serotype of this genus that is based on DNA molecular typing methods.() Typing methods must be simple, rapid, with great repetition and distinction power; however, current methods such as multi locus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) are not only time-consuming, but also low repeatability.()The spa typing is a typing method in Staphylococcus aureus, which is founded on DNA sequencing of short sequence …show more content…
HRM was performed in 20µL reaction contained 4µL of 5x HOT FIRE Pol Eva Green HRM Mix (ROX) (Solis BioDyneCo, Estonia), 1µL of each primer(10pM), 1µL of DNA template (20ng) and 13µL of DNase and RNase free water (SinaClon BioScience Co, Iran). The PCR condition was: 95 °C for10 min, 45 cycles at 95 °C for 30 s, 63 °C for 30s and 72 °C for 30 s. Then, 95 °C for 1 min and 40 °C for 1 min, followed by HRM ramping from 70–95 °C with fluorescence data acquisition at 0.05 °C increments. The spa types t030, t459 and t016100 were used as controls in HRM experiment. The melting curves of all samples were analyzed by High Resolution Melt Software v3.0(Thermo Fisher ScientificCo,
Staphylococcus aureus is one of the most successful human pathogens, which can be detected in varied parts of the world . ()Some factors including virulence factor and drug resistance are efficacious in increasing infectious related death rate. Firstly, Methicillin-resistant Staphylococcus aureus (MRSA) strains were recognized in the early 1960s, moreover, they considered as a nosocomial deathful pathogen due to their role in increasing costs and bedridden length() In 2010 almost 15-20 % of patients with Staphylococcus aureus bacteremia were dead.() One of the most important ways in controlling and preventing of hospital infections is discerning serotype of this genus that is based on DNA molecular typing methods.() Typing methods must be simple, rapid, with great repetition and distinction power; however, current methods such as multi locus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) are not only time-consuming, but also low repeatability.()The spa typing is a typing method in Staphylococcus aureus, which is founded on DNA sequencing of short sequence …show more content…
HRM was performed in 20µL reaction contained 4µL of 5x HOT FIRE Pol Eva Green HRM Mix (ROX) (Solis BioDyneCo, Estonia), 1µL of each primer(10pM), 1µL of DNA template (20ng) and 13µL of DNase and RNase free water (SinaClon BioScience Co, Iran). The PCR condition was: 95 °C for10 min, 45 cycles at 95 °C for 30 s, 63 °C for 30s and 72 °C for 30 s. Then, 95 °C for 1 min and 40 °C for 1 min, followed by HRM ramping from 70–95 °C with fluorescence data acquisition at 0.05 °C increments. The spa types t030, t459 and t016100 were used as controls in HRM experiment. The melting curves of all samples were analyzed by High Resolution Melt Software v3.0(Thermo Fisher ScientificCo,