winnowing dna for rare sequences highly specific sequence and methylation based enrichment筛选dna为罕见的序列高度特定的序列和基于甲基化的浓缩.pdf

Winnowing DNA for Rare Sequences: Highly Specific Sequence and Methylation Based Enrichment 1 2 2 2 1,2 Jason D. Thompson , Gosuke Shibahara , Sweta Rajan , Joel Pel , Andre Marziali * 1 Department of Physics and Astronomy, University of British Columbia, Vancouver, British Columbia, Canada, 2 Boreal Genomics Inc., Vancouver, British Columbia, Canada Abstract Rare mutations in cell populations are known to be hallmarks of many diseases and cancers. Similarly, differential DNA methylation patterns arise in rare cell populations with diagnostic potential such as fetal cells circulating in maternal blood. Unfortunately, the frequency of alleles with diagnostic potential, relative to wild-type background sequence, is often well below the frequency of errors in currently available methods for sequence analysis, including very high throughput DNA sequencing. We demonstrate a DNA preparation and purification method that through non-linear electrophoretic separation in media containing oligonucleotide probes, achieves 10,000 fold enrichment of target DNA with single nucleotide specificity, and 100 fold enrichment of unmodified methylated DNA differing from the background by the methylation of a single cytosine residue. Citation: Thompson JD, Shibahara G, Rajan S, Pel J, Marziali A (2012) Winnowing DNA for Rare Sequences: Highly Specific Sequence and Methylation Based Enrichment. PLoS ONE 7(2): e31597. doi:10.1371/journal.pone.0031597 Editor: Guoliang Xu, Chinese Academy of Science, China Received August 27, 2011; Accepted January 11, 2012; Published February 15, 2012 Copyright: 2012 Thom