ArticlePloS one2016
MeltMan: Optimization, Evaluation, and Universal Application of a qPCR System Integrating the TaqMan qPCR and Melting Analysis into a Single Assay.
Article in PloS one, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. An erratum has been issued. Cited by 9 papers.
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Who cites it
9 citing papers in PubMed.
- Uncovering Molecular Quencher Effects on FRET Phenomena in Microsphere-Immobilized Probe Systems.Analytical chemistry · 2023Article
- Investigation of respiratory disease outbreaks of poultry in Bangladesh using two real-time PCR-based simultaneous detection assays.Frontiers in veterinary science · 2022Article
- Article
- Application of Short Pre-enrichment, and Double Chemistry Real-Time PCR, Combining Fluorescent Probes and an Intercalating Dye, for Same-Day Detection and Confirmation ofFrontiers in microbiology · 2020Article
- Circulation of influenza A and B in the Czech Republic from 2000-2001 to 2015-2016.BMC infectious diseases · 2019Article
- In silico re-assessment of a diagnostic RT-qPCR assay for universal detection of Influenza A viruses.Scientific reports · 2019Article
- Article
- Evaluation of TaqMan qPCR System Integrating Two Identically Labelled Hydrolysis Probes in Single Assay.Scientific reports · 2017Article
- Riems influenza a typing array (RITA): An RT-qPCR-based low density array for subtyping avian and mammalian influenza a viruses.Scientific reports · 2016Article
Corrections and comments
- Erratum issued
Authors and funding
10 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
In the present work, we optimised and evaluated a qPCR system integrating 6-FAM (6-carboxyfluorescein)-labelled TaqMan probes and melting analysis using the SYTO 82 (S82) DNA binding dye in a single reaction. We investigated the influence of the S82 on various TaqMan and melting analysis parameters and defined its optimal concentration. In the next step, the method was evaluated in 36 different TaqMan assays with a total of 729 paired reactions using various DNA and RNA templates, including field specimens. In addition, the melting profiles of interest were correlated with the electrophoretic patterns. We proved that the S82 is fully compatible with the FAM-TaqMan system. Further, the advantages of this approach in routine diagnostic TaqMan qPCR were illustrated with practical examples. These included solving problems with flat or other atypical amplification curves or even false negativity as a result of probe binding failure. Our data clearly show that the integration of the TaqMan qPCR and melting analysis into a single assay provides an additional control option as well as the opportunity to perform more complex analyses, get more data from the reactions, and obtain analysis results with higher confidence.
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