Winnowing DNA for rare sequences: highly specific sequence and methylation based enrichment

PLoS One. 2012;7(2):e31597. doi: 10.1371/journal.pone.0031597. Epub 2012 Feb 15.


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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • CpG Islands / genetics*
  • Cytosine / chemistry*
  • DNA / analysis*
  • DNA / genetics*
  • DNA / isolation & purification
  • DNA Methylation*
  • DNA-Binding Proteins / genetics
  • Enhancer of Zeste Homolog 2 Protein
  • Humans
  • Mutation / genetics
  • Oligonucleotide Probes*
  • Polycomb Repressive Complex 2
  • Real-Time Polymerase Chain Reaction
  • Transcription Factors / genetics


  • DNA-Binding Proteins
  • Oligonucleotide Probes
  • Transcription Factors
  • Cytosine
  • DNA
  • EZH2 protein, human
  • Enhancer of Zeste Homolog 2 Protein
  • Polycomb Repressive Complex 2