Broad-Range Ribosomal RNA Real-Time PCR after Removal of DNA from Reagents: Melting Profiles for Clinically Important Bacteria

Tseng, Ching Ping; Cheng, Ju Chien; Tseng, Ching Chung; Wang, Cheyien; Chen, Yi Ling; Chiu, Daniel; Liao, How Chin; Chang, Shy Shin

doi:10.1373/49.2.306

Cited by 33 publications

(23 citation statements)

References 24 publications

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“…The crossing points for the broad-range real-time PCR ranged from [9.52 (0.34), n ϭ 4, to 27.88 (0.99), n ϭ 4] for 100 ng to 100 fg of E. coli genomic DNA. For the no-template control (NTC) 9 reaction, the crossing point was at the cycle number of [28.20 (0.85…”

Section: Resultsmentioning

confidence: 99%

“…1B in the online Data Supplement). Because of the presence of contaminated bacterial DNA in the PCR reagents (9,18,19 ), a small melting peak was observed for NTC. Low concentrations of initial template DNA usually led to a primer dimer with a melting peak at ϳ74°C.…”

Section: Resultsmentioning

confidence: 99%

“…For example, denaturing HPLC has been used for rapid detection and identification of various bacterial species (20 ). SYBR Green I-based broad-range real-time PCR followed by melting curve analysis has also been used to enhance the speed of bacterial detection (9 ). A recent study elucidated the application of high-resolution melting analysis of heat shock protein 65 PCR products for rapid species identification within the Mycobacterium chelonae-abscessus group (21 ).…”

Section: Discussionmentioning

confidence: 99%

“…For rapid detection of bacteria, we used the primer pair p201 and p1370 for real-time amplification of 16S rRNA gene, with PCR product lengths of 216 or 217 bp (9 ). The 176-bp interprimer region likely contains information for at least partial phylogenetic characterization for subclassifying clinically important bacterial species (10 ).…”

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confidence: 99%

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Rapid Detection and Identification of Clinically Important Bacteria by High-Resolution Melting Analysis after Broad-Range Ribosomal RNA Real-Time PCR

Huang²,

et al. 2006

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Background: Broad-range PCR provides valuable information for detecting bacterial infections. This study assesses the combined use of broad-range real-time PCR and high-resolution melting analysis for rapid detection and identification of clinically important bacteria. Methods: We subjected 46 bacterial culture colonies representing 25 clinically important bacterial species to LightCycler real-time PCR amplification of the 16S rRNA gene in the presence of LCGreen I fluorescent dye. We performed high-resolution melting analysis of the PCR products with the HR-1 instrument and used melting profiles as molecular fingerprints for bacterial species identification. We validated this method via assessment of 54 consecutive bacteria culture colonies obtained from a clinical microbiology laboratory. Results: The 16S rRNA gene of all 25 bacterial species was amplifiable by this method, with PCR product

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Rapid Detection and Identification of Clinically Important Bacteria by High-Resolution Melting Analysis after Broad-Range Ribosomal RNA Real-Time PCR

Huang²,

et al. 2006

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“…Real-time PCR was carried out by using the QuantiTect SYBR green PCR kit (Qiagen) and a LightCycler LC480 thermocycler (Roche). After PCR, the cycle threshold (C T ) values were normalized to an average C T value of amplifications (⌬C T ) performed with 2 different universal primer pairs for the domain Bacteria (7,8). The relative amount of each taxon was finally calculated as 2…”

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confidence: 99%

Influence of Salmonella enterica Serovar Enteritidis Infection on the Development of the Cecum Microbiota in Newly Hatched Chicks

Juřicová

Vídeňská

Lukač

et al. 2013

Appl Environ Microbiol

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Terminal restriction fragment length polymorphism and quantitative PCR showed that the cecal microbiota of chicks up to the age of 21 days was dominated by representatives of the orders Enterobacteriales, Clostridiales, and Lactobacillales. Salmonella enterica serovar Enteritidis infection caused the greatest changes in the gut microbiota when 1-day-old chicks were infected, compared with the infection of 4-and 16-day-old chicks.

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Oil production by a consortium of oleaginous microorganisms grown on primary effluent wastewater

Hall

Hetrick

French

et al. 2010

J of Chemical Tech & Biotech

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Broad-Range Ribosomal RNA Real-Time PCR after Removal of DNA from Reagents: Melting Profiles for Clinically Important Bacteria

Cited by 33 publications

References 24 publications

Rapid Detection and Identification of Clinically Important Bacteria by High-Resolution Melting Analysis after Broad-Range Ribosomal RNA Real-Time PCR

Rapid Detection and Identification of Clinically Important Bacteria by High-Resolution Melting Analysis after Broad-Range Ribosomal RNA Real-Time PCR

Influence of Salmonella enterica Serovar Enteritidis Infection on the Development of the Cecum Microbiota in Newly Hatched Chicks

Oil production by a consortium of oleaginous microorganisms grown on primary effluent wastewater

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