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BioMed Research International
Volume 2014, Article ID 641090, 11 pages
Research Article

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

1Research and Development Department, Thorne Diagnostics Inc. 100 Cummings Center, Beverly, MA 01915, USA
2Department of Pathology, Icahn School of Medicine at Mount Sinai, 1428 Madison Avenue, New York City, NY 10029, USA

Received 12 August 2013; Accepted 17 January 2014; Published 2 March 2014

Academic Editor: Michael Kalafatis

Copyright © 2014 James H. Smith et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.


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.