http://www.nature.com/nprot/jour ... nprot.2011.444.html ( U) _+ Y8 i" i$ D8 `3 I4 b 7 t B2 V- P$ |Nat Protoc. 2012 Jan 19;7(2):256-67. doi: 10.1038/nprot.2011.444. . q4 m6 c& B8 P. ]; H/ s; w5 BUsing formaldehyde-assisted isolation of regulatory elements (FAIRE) to isolate active regulatory DNA. / r/ e4 N( V$ `( P4 \Simon JM, Giresi PG, Davis IJ, Lieb JD. C/ X- b4 z3 G o9 I* _
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Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.9 {* z u: Y$ o% ]1 D3 K# d
6 e- F" F+ i% R" M7 Z+ z0 e$ {Abstract3 M9 J! {& b- v4 T$ @- h0 V; H* ]7 A
Eviction or destabilization of nucleosomes from chromatin is a hallmark of functional regulatory elements in eukaryotic genomes. Historically identified by nuclease hypersensitivity, these regulatory elements are typically bound by transcription factors or other regulatory proteins. FAIRE (formaldehyde-assisted isolation of regulatory elements) is an alternative approach to identify these genomic regions and has proven successful in a multitude of eukaryotic cell and tissue types. Cells or dissociated tissues are cross-linked briefly with formaldehyde, lysed and sonicated. Sheared chromatin is subjected to phenol/chloroform extraction and the isolated DNA, typically encompassing 1-3% of the human genome, is purified. We provide guidelines for quantitative analysis by PCR, microarrays or next-generation sequencing. Regulatory elements enriched by FAIRE have high concordance with those identified by nuclease hypersensitivity or chromatin immunoprecipitation (ChIP), and the entire procedure can be completed in 3 d. FAIRE has low technical variability, which allows its usage in large-scale studies of chromatin from normal or diseased tissues.作者: hzh0525 时间: 2012-7-14 08:18