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Differential roles of promoters, enhancers, and insulators in cell-type specific gene expression

ABSTRACT: Chromatin immunoprecipitation was performed using antibodies against CTCF and formaldehyde crosslinked chromatin from K562 Chromatin cells. The immunoprecipitated DNA was amplified, fluorescently labeled and hybridized to a 391K element tiling array based on NCBI35 build of the human genome (hg17) that contains overlapping 50-mer oligonucleotides, positioned at every 38 basepairs (bp) throughout the ENCODE regions. Keywords: ChIP-chip

ORGANISM(S): Homo sapiens

PROVIDER: GSE8098 | GEO | 2008/11/01

SECONDARY ACCESSION(S): PRJNA100953

REPOSITORIES: GEO

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Differential roles of promoters, enhancers, and insulators in cell-type specific gene expression

Project description:Chromatin immunoprecipitation was performed using antibodies against CTCF and formaldehyde crosslinked chromatin from K562 Chromatin cells. The immunoprecipitated DNA was amplified, fluorescently labeled and hybridized to a 391K element tiling array based on NCBI35 build of the human genome (hg17) that contains overlapping 50-mer oligonucleotides, positioned at every 38 basepairs (bp) throughout the ENCODE regions. Keywords: ChIP-chip 3 ChIP-chip Samples

2010-05-17 | E-GEOD-8098 | biostudies-arrayexpress

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FAIRE performed in human foreskin fibroblast cells

Project description:To perform FAIRE, chromatin is crosslinked with formaldehyde in vivo, sheared by sonication, and phenol-chloroform extracted. The DNA recovered in the aqueous phase is fluorescently labeled and hybridized to a DNA microarray. FAIRE performed in human cells strongly enriches DNA coincident with the location of DNaseI hypersensitive sites, transcriptional start sites, and active promoters. Keywords: Genome wide map of active chromatin

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Project description:This data includes regulatory factor profiling using ChIP-seq. Cells were grown according to the approved ENCODE cell culture protocols. Cells were crosslinked with 1% formaldehyde, and the reaction was quenched by the addition of glycine. Fixed cells were rinsed with PBS, lysed in nuclei lysis buffer, and the chromatin was sheared to 200-500 bp fragments using Fisher Dismembrator (model 500). Sheared chromatin fragments were immunoprecipitated with specific polyclonal antibodies at 4 degrees C with gentle rotation. Antibody-chromatin complexes were washed and eluted. The cross linking in immunoprecipitated DNA was reversed and treated with RNase-A. Following proteinase K treatment, the DNA fragments were purified by phenol-chloroform-isoamyl alcohol extraction and ethanol precipitation. 20-50 ng of ChIP DNA was end-repaired, adenine ligated to Illumina adapters was added, and then a Solexa library was made for sequencing. ChIP-seq affinity is directly reflected in raw tag density (Raw Signal), which is shown in the track as density of tags mapping within a 150 bp sliding window (at a 20 bp step across the genome). ChIP-seq affinity zones (HotSpots) were identified using the HotSpot algorithm described in Sabo et al. (2004). 1.0% false discovery rate thresholds (FDR 0.01) were computed for each cell type by applying the HotSpot algorithm to an equivalent number of random uniquely mapping 36mers. ChIP-seq affinity (Peaks) were identified as signal peaks within FDR 1.0% hypersensitive zones using a peak-finding algorithm.

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H3K27me3 ChIP-seq in DNMT triple knock-out mESCs

Project description:We immunoprecipitated formaldehyde crosslinked chromatin from DNMT triple knockout cells using an antibody against methylated lysine 27 on histone H3. Subsequent high-throughput sequencing allowed us to study the genome-wide distribution of this histone mark in the absence of DNA methylation.

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Project description:Maize earshoot MOA-seq; open chromatin profiling using micrococcal nuclease with formaldehyde-crosslinked nuclei.

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Project description:In crosslinked MINUTE-ChIP, formaldehyde-fixed chromatin is fragmented using sonication, blunted and ligated to double-stranded DNA adaptors that include a T7 RNA polymerase promoter and a sample barcode sequence. Finally, samples are combined and subsequent ChIP reactions are performed with the pooled samples. ChIP material is prepared into an Illumina-compatible library using linear amplification by virtue of T7 RNA polymerase, reverse transcription and a low-cycle library PCR amplification. Here, we demonstrate a MINUTE-ChIP for Nanog from formaldehyde-fixed cells using fragmentation by sonication.

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