ArticleNucleic acids research2013
Combination of fluorescence color and melting temperature as a two-dimensional label for homogeneous multiplex PCR detection.
Article in Nucleic acids research, 2013. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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Who cites it
23 citing papers in PubMed.
- A Novel Molecular Method for Simultaneous Identification ofFrontiers in cellular and infection microbiology · 2021Trial
- Universal primer mediated multiplex asymmetric PCR coupled with hybridization-based electrochemical biosensing for rapid, simultaneous and label-free detection of multiple pathogens.Mikrochimica acta · 2026Article
- Application of Universal LNA Probe-Mediated Multiplex Real-Time PCR for the Detection of Antibiotic Resistance Genes.Molecular biotechnology · 2026Article
- Robust dPCR Hyper-Plexing Approach by Color Combination for Detection of 17 ESR1 Mutations in Breast Cancer Patient Samples.Methods in molecular biology (Clifton, N.J.) · 2026Article
- An Efficient Pharmacogenomic Assay for Psychotropic Drugs by Multiplex Fluorescence Melting Curve Analysis.Drug design, development and therapy · 2026Article
- Cas12a Cis-cleavage mediated lateral flow assay enables multiplex and ultra-specific nucleic acid detection.Nature communications · 2025Article
- Advancing quantitative PCR with color cycle multiplex amplification.Nucleic acids research · 2024Article
- Rapid and specific differentiation of Salmonella enterica serotypes typhi and Paratyphi by multicolor melting curve analysis.Gut pathogens · 2024Article
- Highly multiplex PCR assays by coupling the 5'-flap endonuclease activity ofProceedings of the National Academy of Sciences of the United States of America · 2022Article
- PlexProbes enhance qPCR multiplexing by discriminating multiple targets in each fluorescent channel.PloS one · 2022Article
- Single-tube detection of nine bacterial antibiotic-resistance genes by a 2-dimensional multiplex qPCR assay based on fluorescence and melting temperature.Molecular biology reports · 2020Article
- Simultaneous Identification of Clinically CommonFrontiers in cellular and infection microbiology · 2019Article
- Rapid Identification of Clinically RelevantJournal of clinical microbiology · 2019Article
- Evaluation of Real-Time PCR Coupled With Multiplex Probe Melting Curve Analysis for Pathogen Detection in Patients With Suspected Bloodstream Infections.Frontiers in cellular and infection microbiology · 2019Article
- A novel multiplex PCR for virus detection by melting curve analysis.Journal of virological methods · 2018Article
- Ratiometric Fluorescence Coding for Multiplex Nucleic Acid Amplification Testing.Analytical chemistry · 2018Article
- Article
- Establishment of a Gene Detection System for Hotspot Mutations of Hearing Loss.BioMed research international · 2018Article
- Simultaneous Identification of Ten Bacterial Pathogens Using the Multiplex Ligation Reaction Based on the Probe Melting Curve Analysis.Scientific reports · 2017Article
- Spoligotyping of Mycobacterium tuberculosis Complex Isolates by Use of Ligation-Based Amplification and Melting Curve Analysis.Journal of clinical microbiology · 2016Article
Corrections and comments
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Authors and funding
6 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Multiplex analytical systems that allow detection of multiple nucleic acid targets in one assay can provide rapid characterization of a sample while still saving cost and resources. However, few systems have proven to offer a solution for mid-plex (e.g. 10- to 50-plex) analysis that is high throughput and cost effective. Here we describe the combined use of fluorescence color and melting temperature (Tm) as a virtual 2D label that enables homogenous detection of one order of magnitude more targets than current strategies on real-time polymerase chain reaction platform. The target was first hybridized with a pair of ligation oligonucleotides, one of which harbored an artificial sequence that had a unique Tm when hybridized with a reporter fluorogenic probe. The ligated products were then amplified by a universal primer pair and denatured by a melting curve analysis procedure. The targets were identified by their respective Tm values in the corresponding fluorescence detection channels. The proof-of-principle of this approach was validated by genotyping 15 high-risk human papillomaviruses and 48 human single-nucleotide polymorphisms. The robustness of this method was demonstrated by analyzing a large number of clinical samples in both cases. The combined merits of multiplexity, flexibility and simplicity should make this approach suitable for a variety of applications.
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