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Figure FAIRE Procedure (A) The FAIRE procedure described in the text is shown on the left, while preparation of the reference or input sample is shown on the right. The DNA recovered from he aqueous phase of each extraction can then be used to identify sites of open chromatin using qPCR, tiling microarrays, or high-throughput sequencing applications. (B) For qPCR, a series of primers, depicted as convergent arrows, are designed to span a genomic region of interest. Sites of open chromatin are highlighted in blue, with qPCR results depicted above. Amplicons that span or are near the boundaries of open chromatin often result in lower relative enrichment due to shearing of DNA fragments, as shown by asterisks. (C) Microarrays. Typically we use highresolution microarrays that tile either regions of interest or the entire genome of an organism with 50 to 70 bp oligonucleotides. (D) High-throughput sequencing technologies can be used to map the DNA fragments back to the reference genome.【Giresi P G, Lieb J D. Isolation of active regulatory elements from eukaryotic chromatin using FAIRE (Formaldehyde Assisted Isolation of Regulatory Elements)[J]. Methods, 2009, 48(3):233-239.】

Comparison of experimental protocols.Experiments to detect different aspects of DNA-binding proteins share many of the same steps; simplified schematics of the main steps are shown. a | Chromatin immunoprecipitation followed by sequencing (ChIP–seq) for DNA-binding proteins such as transcription factors. Recent variations on the standard protocol include using endonuclease digestion instead of sonication (ChIP–exo) to increase the resolution of binding-site detection and to eliminate contaminating DNA, and DNA amplification after ChIP for samples with limited cells. b | ChIP–seq for histone modifications uses micrococcal nuclease (MNase) digestion to fragment DNA and can also now be run on low-quantity samples when combined with the additional post-ChIP amplification. c | DNase–seq relies on digestion by the DNaseI nuclease to identify regions of nucleosome-depleted open chromatin where there are binding sites for all types of factors, but it cannot identify what specific factors are bound. d | Formaldehyde-assisted identification of regulatory elements (FAIRE–seq) similarly identifies nucleosome-depleted regions by extracting fragmented DNA that is not crosslinked to nucleosomes. LinDA, single-tube linear DNA amplification; T7, T7 phage RNA polymerase.【Furey, Terrence S . ChIP–seq and beyond: new and improved methodologies to detect and characterize protein–DNA interactions[J]. Nature Reviews Genetics, 2012, 13(12):840-852.】