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Nucleic Acids Research 2006 34(3):e17; doi:10.1093/nar/gnj017
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Published online 7 February 2006

© The Author 2006. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oxfordjournals.org

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Accurate quantification of DNA methylation using combined bisulfite restriction analysis coupled with the Agilent 2100 Bioanalyzer platform

Romulo Martin Brena1,5, Herbert Auer2, Karl Kornacker3, Björn Hackanson5,6, Aparna Raval5, John C. Byrd4 and Christoph Plass5,*

1Department of Molecular Genetics, The Ohio State University Columbus, OH, USA 2Columbus Children's Research Institute Columbus, OH, USA 3Division of Sensory Biophysics, The Ohio State University Columbus, OH, USA 4Department of Medicine and the Comprehensive Cancer Center, Divisions of Hematology-Oncology, The Ohio State University Columbus, OH, USA 5Department of Molecular Virology, Immunology and Medical Genetics, The Ohio State University Columbus, OH, USA 6Department of Hematology, University of Freiburg Medical Center Freiburg, Germany

*To whom correspondence should be addressed at Division of Human Cancer Genetics, Medical Research Facility Room 464A, 420 West 12th Avenue, Columbus, OH 43210, USA. Tel: +1 614 292 6505; Fax: +1 614 688 4761; Email: christoph.plass{at}osumc.edu

Received November 7, 2005. Revised January 19, 2006. Accepted January 19, 2006.

DNA methylation is the best-studied epigenetic modification and describes the conversion of cytosine to 5-methylcytosine. The importance of this phenomenon is that aberrant promoter hypermethylation is a common occurrence in cancer and is frequently associated with gene silencing. Various techniques are currently available for the analysis of DNA methylation. However, accurate and reproducible quantification of DNA methylation remains challenging. In this report, we describe Bio-COBRA (combined bisulfite restriction analysis coupled with the Agilent 2100 Bioanalyzer platform), as a novel approach to quantitative DNA methylation analysis. The combination of a well-established method, COBRA, which interrogates DNA methylation via the restriction enzyme analysis of PCR-amplified bisulfite treated DNAs, with the Bioanalyzer platform allows for the rapid and quantitative assessment of DNA methylation patterns in large sample sets. The sensitivity and reproducibility of Bio-COBRA make it a valuable tool for the analysis of DNA methylation in clinical samples, which could aid in the development of diagnostic and prognostic parameters with respect to disease detection and management.


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