DNase I Digestion of Isolated Nulcei for Genome-Wide Mapping of DNase Hypersensitivity Sites in Chromatin
DNase I hypersensitivity (DHS) analysis is a powerful method to analyze chromatin structure and identify genomic regulatory elements. Integration of a high-throughput detection method into DHS analysis makes genome-wide mapping of DHS sites possible at a reasonable cost. Here we describe methods for DHS analysis carried out with mouse liver nuclei, involving DNase I digestion followed by isolation of DNase I-released DNA fragments suitable for high-throughput, next generation DNA sequencing (DNase-seq). A real-time PCR-based assay used to optimize DNase I digestion conditions is also described.
Key wordsDNase I hypersensitivity assay Next generation DNA sequencing DNase-seq Chromatin structure
Supported in part by NIH grant DK33765 (to DJW).
- 6.Sabo PJ, Kuehn MS, Thurman R, Johnson BE, Johnson EM, Cao H, Yu M, Rosenzweig E, Goldy J, Haydock A, Weaver M, Shafer A, Lee K, Neri F, Humbert R, Singer MA, Richmond TA, Dorschner MO, McArthur M, Hawrylycz M, Green RD, Navas PA, Noble WS, Stamatoyannopoulos JA (2006) Genome-scale mapping of DNase I sensitivity in vivo using tiling DNA microarrays. Nat Methods 3:511–518PubMedCrossRefGoogle Scholar
- 10.Weis JH, Quertermous T (2001) Size fractionation using sucrose gradients. Curr Protoc Mol Biol, Chapter 5:Unit 5.3Google Scholar