Detection of Genetic Variations in Coagulopathy-Related Genes Using Ramified Rolling Circle Amplification

We evaluated single nucleotide polymorphism (SNP) detection via a target-capture, C-probe ligation, and RAM assay in a single-blind comparison to clinical samples that had been tested with FDA-cleared tests for up to 4 different vascular disease-related SNPs. In the RAM assay circulizable linear pro...

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Bibliographic Details
Published in:BioMed research international 2014-01, Vol.2014 (2014), p.1-11
Main Authors: Smith, James H., Cui, Miao, Zhang, David Y., Beals, Thomas P., Ye, Fei
Format: Article
Language:English
Subjects:
Automation
Cardiovascular disease
Deoxyribonucleic acid
DNA
DNA - chemistry
DNA Primers - chemistry
DNA Probes - chemistry
Enzymes
Genes
Genetic testing
Hybridization
Laboratories
Mutation
Noise
Nucleic Acid Amplification Techniques - methods
Polymorphism, Single Nucleotide
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Summary:We evaluated single nucleotide polymorphism (SNP) detection via a target-capture, C-probe ligation, and RAM assay in a single-blind comparison to clinical samples that had been tested with FDA-cleared tests for up to 4 different vascular disease-related SNPs. In the RAM assay circulizable linear probes (C- or padlock probes) were annealed directly to genomic DNA, processed on a largely automated platform, and ligated C-probes were amplified by real-time RAM. After allele determinations were made with the experimental system, the sample genotypes were unblinded and the experimentally determined genotypes were found to be completely consistent with the FDA-cleared test results. The methods and results presented here show that a combination of C-probes, automated sample processing, and isothermal RAM provides a robust, and specific, nucleic acid detection platform that is compatible with automated DNA sample preparation and the throughput requirements of the clinical laboratory.
ISSN:2314-6133
2314-6141
DOI:10.1155/2014/641090