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1860799948267716608
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INTELEK Repository
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Online Access
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https://intelek.unisza.edu.my/intelek/pages/search.php?search=!collection407072
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2024-09-24 17:54
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Restricted Document
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8033
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UniSZA
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K Hiramatsu . Vancomycin-resistance Staphylococcus aureus: a new model of antibiotic resistance. Lancet Infect. Dis. 200 1; 1, 147-55. FJ Schmitz, AC Fluit, M Luckefah, B Engler, B Hofmann, J Verhoef, HP Heinz, U Hadding and ME Jone. The effect of reserpine, an inhibitor of multidrug efflux pumps, on the in-vitro activities of ciprofloxacin, 27 sparfloxacin and moxifloxacin against clinical isolates of Staphylococcus aureus. J . Antimicrob. Chemother. 1998; 42, 807-10 . TY Tan . Use of molecular techniques for the detection of antibiotic resistance in bacteria . Expet. Rev. Mol. Diagn. 2003; 3, 93-103. CJ Papasian and 8 Garrison . Evaluation of a rapid slide agglutination test for identification of Staphylococcus aureus. Diagn . Microbiol . Infect. Dis. 1999; 33, 201-3. MB Miller, H Meyer, E Rogers and PH Gilligan. Comparison of conventional susceptibility testing, penicillin-binding protein 2a latex agglutination testing, and mecA real-time PCR for detection of oxacillin resistance in Staphylococcus aureus and coagulase-negative Staphylococcus. J. Clin. Microbiol.2005; 43,
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oai_dc
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https://intelek.unisza.edu.my/intelek/pages/view.php?ref=8033
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8033 https://intelek.unisza.edu.my/intelek/pages/view.php?ref=8033 https://intelek.unisza.edu.my/intelek/pages/search.php?search=!collection407072 Restricted Document Article Journal Deflate/Inflate 900 1440x900 1440 image/png 2024-09-24 17:54 3891-01-FH02-FPBSM-14-00436.png UniSZA Private Access In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates Walailak Journal of Science and Technology (WJST) The aims of this study were to validate the primers developed for molecular-based detection and identification of Malaysian clinical isolates of methicilin-resistance Staphylococcus aureus (MRSA) using in-silico Polymerase Chain Reaction (PCR) and real-time PCR SYBR with Green I. Rapid molecular diagnostic and risk assessment of the MRSA are possible by real-time PCR SYBR Green I. However, validation of such primers for real-life samples is expensive and time consuming. Hence, development and verification of real-time PCR primers by in-silico PCR can be the first step in the selection of the most appropriate primers. Three species-specific markers were chosen targeting coa (staphylocoagulase), nuc (thermonuclease) and mecA (methicillin-resistance) and were specifically verified against 35 selected S. aureus strains by using in-silico PCR. For the actual laboratory verification, all of the 3 genes were detected with a single specific melting curve peak (Tm at 76.16 ± 0.8 °C, 78.50 ± 0.4 °C and 74.41 ± 0.6 °C for a coa, nuc and mecA respectively) in 32 bacterial strains including ATCC reference strains. Thus, there is no disagreement between both in-silico PCR and real-time PCR verification and validation of the primers designed for the detection and identification of MRSA in this study. The potential of using a bioinformatics approach (in-silico PCR) before selecting primer pairs for a given study may enable researchers to accept or reject the potential primer pairs for downstream experimental (in vitro) PCR without wasting any chemicals as well as related cost. 10 3 237-246 K Hiramatsu . Vancomycin-resistance Staphylococcus aureus: a new model of antibiotic resistance. Lancet Infect. Dis. 200 1; 1, 147-55. FJ Schmitz, AC Fluit, M Luckefah, B Engler, B Hofmann, J Verhoef, HP Heinz, U Hadding and ME Jone. The effect of reserpine, an inhibitor of multidrug efflux pumps, on the in-vitro activities of ciprofloxacin, 27 sparfloxacin and moxifloxacin against clinical isolates of Staphylococcus aureus. J . Antimicrob. Chemother. 1998; 42, 807-10 . TY Tan . Use of molecular techniques for the detection of antibiotic resistance in bacteria . Expet. Rev. Mol. Diagn. 2003; 3, 93-103. CJ Papasian and 8 Garrison . Evaluation of a rapid slide agglutination test for identification of Staphylococcus aureus. Diagn . Microbiol . Infect. Dis. 1999; 33, 201-3. MB Miller, H Meyer, E Rogers and PH Gilligan. Comparison of conventional susceptibility testing, penicillin-binding protein 2a latex agglutination testing, and mecA real-time PCR for detection of oxacillin resistance in Staphylococcus aureus and coagulase-negative Staphylococcus. J. Clin. Microbiol.2005; 43,
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| spellingShingle |
In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates
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| summary |
The aims of this study were to validate the primers developed for molecular-based detection and identification of Malaysian clinical isolates of methicilin-resistance Staphylococcus aureus (MRSA) using in-silico Polymerase Chain Reaction (PCR) and real-time PCR SYBR with Green I. Rapid molecular diagnostic and risk assessment of the MRSA are possible by real-time PCR SYBR Green I. However, validation of such primers for real-life samples is expensive and time consuming. Hence, development and verification of real-time PCR primers by in-silico PCR can be the first step in the selection of the most appropriate primers. Three species-specific markers were chosen targeting coa (staphylocoagulase), nuc (thermonuclease) and mecA (methicillin-resistance) and were specifically verified against 35 selected S. aureus strains by using in-silico PCR. For the actual laboratory verification, all of the 3 genes were detected with a single specific melting curve peak (Tm at 76.16 ± 0.8 °C, 78.50 ± 0.4 °C and 74.41 ± 0.6 °C for a coa, nuc and mecA respectively) in 32 bacterial strains including ATCC reference strains. Thus, there is no disagreement between both in-silico PCR and real-time PCR verification and validation of the primers designed for the detection and identification of MRSA in this study. The potential of using a bioinformatics approach (in-silico PCR) before selecting primer pairs for a given study may enable researchers to accept or reject the potential primer pairs for downstream experimental (in vitro) PCR without wasting any chemicals as well as related cost.
|
| title |
In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates
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| title_full |
In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates
|
| title_fullStr |
In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates
|
| title_full_unstemmed |
In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates
|
| title_short |
In silico PCR Verification and Simplex Real-Time PCR Detection of Methicillin-Resistant Staphylococcus aureus (MRSA) from East Coast Malaysian Clinical Isolates
|
| title_sort |
in silico pcr verification and simplex real-time pcr detection of methicillin-resistant staphylococcus aureus (mrsa) from east coast malaysian clinical isolates
|