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Publication Detail
Methylome analysis using MeDIP-seq with low DNA concentrations.
  • Publication Type:
    Journal article
  • Publication Sub Type:
    Journal Article
  • Authors:
    Taiwo O, Wilson GA, Morris T, Seisenberger S, Reik W, Pearce D, Beck S, Butcher LM
  • Publication date:
    04/2012
  • Pagination:
    617, 636
  • Journal:
    Nat Protoc
  • Volume:
    7
  • Issue:
    4
  • Country:
    England
  • PII:
    nprot.2012.012
  • Language:
    eng
  • Keywords:
    Animals, Bone Marrow Cells, DNA Fragmentation, DNA Methylation, Epigenomics, Flow Cytometry, Gene Library, Immunoprecipitation, Mice, Sequence Analysis, DNA
Abstract
DNA methylation is an epigenetic mark that has a crucial role in many biological processes. To understand the functional consequences of DNA methylation on phenotypic plasticity, a genome-wide analysis should be embraced. This in turn requires a technique that balances accuracy, genome coverage, resolution and cost, yet is low in DNA input in order to minimize the drain on precious samples. Methylated DNA immunoprecipitation-sequencing (MeDIP-seq) fulfils these criteria, combining MeDIP with massively parallel DNA sequencing. Here we report an improved protocol using 100-fold less genomic DNA than that commonly used. We show comparable results for specificity (>97%) and enrichment (>100-fold) over a wide range of DNA concentrations (5,000-50 ng) and demonstrate the utility of the protocol for the generation of methylomes from rare bone marrow cells using 160-300 ng of starting DNA. The protocol described here, i.e., DNA extraction to generation of MeDIP-seq library, can be completed within 3-5 d.
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