ArticleBMC genomics2007
Quantitative multiplex detection of plant pathogens using a novel ligation probe-based system coupled with universal, high-throughput real-time PCR on OpenArrays.
Article in BMC genomics, 2007. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed, 72 citations in OpenAlex.
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- Development and Evaluation of Isothermal Amplification Methods for Rapid Detection of LethalFrontiers in microbiology · 2019Article
- Advanced DNA-Based Point-of-Care Diagnostic Methods for Plant Diseases Detection.Frontiers in plant science · 2017Review
- Molecular inversion probe: a new tool for highly specific detection of plant pathogens.PloS one · 2014Article
- Molecular analysis of meso- and thermophilic microbiota associated with anaerobic biowaste degradation.BMC microbiology · 2012Article
- Molecular techniques for pathogen identification and fungus detection in the environment.IMA fungus · 2011Article
- Emerging molecular assays for detection and characterization of respiratory viruses.Clinics in laboratory medicine · 2009Review
- Accurate quantification of microorganisms in PCR-inhibiting environmental DNA extracts by a novel internal amplification control approach using Biotrove OpenArrays.Applied and environmental microbiology · 2009Article
- Robust detection and identification of multiple oomycetes and fungi in environmental samples by using a novel cleavable padlock probe-based ligation detection assay.Applied and environmental microbiology · 2009Article
- Optimised padlock probe ligation and microarray detection of multiple (non-authorised) GMOs in a single reaction.BMC genomics · 2008Article
- 16S-Ribosomal DNA to diagnose culture-negative endocarditis.Current infectious disease reports · 2008Article
- Development and experimental validation of a predictive threshold cycle equation for quantification of virulence and marker genes by high-throughput nanoliter-volume PCR on the OpenArray platform.Applied and environmental microbiology · 2008Article
Corrections and comments
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Authors and funding
7 authors at 6 institutions in 4 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundDiagnostics and disease-management strategies require technologies to enable the simultaneous detection and quantification of a wide range of pathogenic microorganisms. Most multiplex, quantitative detection methods available suffer from compromises between the level of multiplexing, throughput and accuracy of quantification. Here, we demonstrate the efficacy of a novel, high-throughput, ligation-based assay for simultaneous quantitative detection of multiple plant pathogens. The ligation probes, designated Plant Research International-lock probes (PRI-lock probes), are long oligonucleotides with target complementary regions at their 5' and 3' ends. Upon perfect target hybridization, the PRI-lock probes are circularized via enzymatic ligation, subsequently serving as template for individual, standardized amplification via unique probe-specific primers. Adaptation to OpenArrays, which can accommodate up to 3072 33 nl PCR amplifications, allowed high-throughput real-time quantification. The assay combines the multiplex capabilities and specificity of ligation reactions with high-throughput real-time PCR in the OpenArray, resulting in a flexible, quantitative multiplex diagnostic system.
resultsThe performance of the PRI-lock detection system was demonstrated using 13 probes targeting several significant plant pathogens at different taxonomic levels. All probes specifically detected their corresponding targets and provided perfect discrimination against non-target organisms with very similar ligation target sites. The nucleic acid targets could be reliably quantified over 5 orders of magnitude with a dynamic detection range of more than 104. Pathogen quantification was equally robust in single target versus mixed target assays.
conclusionThis novel assay enables very specific, high-throughput, quantitative detection of multiple pathogens over a wide range of target concentrations and should be easily adaptable for versatile diagnostic purposes.
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