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Sample GSM2251439 Query DataSets for GSM2251439
Status Public on Dec 06, 2016
Title CTCF-HL-E105-Wt-Mm-Rep2-L7327-1
Sample type SRA
Source name Embryonic Limb Bud
Organism Mus musculus
Characteristics tissue: hindlimb
developmental stages: E10.5
strain: CD1
chip antibody: CTCF (Active motif: 613111)
Treatment protocol For chromatin modifications, chromatin was prepared from the different tissues with a 1% FA crosslinking for 15 minutes and resuspended in buffer 3 for sonication (Lee et al.,). For CTCF and RAD21 ChIP-seq we prepared chromatin as follow: tissues were disrupted in 0.1% collagenase at 37°C and homogenized using a needle. Cell were then centrifuged and resuspended in (10%FCS, 0.2%Cs, 1% L-Glu, 0.5% Pen-Strep in DMEM:HAM’s F-12 1:1)) and fixed in 1% FA for 10’ on ice. Cell were then lysed in buffer 1 and 2 and resuspended in buffer 3 for sonication (Lee et al. 2006).
Growth protocol Limb bud and midbrain were micro-dissected from mouse embryos
Extracted molecule genomic DNA
Extraction protocol Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with antibody.
We sheared chromatin using Bioruptor until reaching a fragment size of 200-500bp. 10-15μg of chromatin was then used for each replicate chromatin modification ChIP and 30μg for CTCF and RAD21 ChIP. ChIP for H3K4me1 (Abcam: 8898), H3K4me3 (Milipore: 07-473), H3K27Ac (Diagenode: C1540174), H3K27me3 (Milipore: 07-449), CTCF (Active motif: 613111) and RAD21 (Abcam: ab992) was then performed as in Lee et al., 2006. Libraries were prepared using the Nextera adaptors and sequenced.
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 2500
Description The merged .bw file for this sample is available on GSM2251438
Data processing Single-end reads from ChIP-seq experiments were mapped with Bowtie-2.2.6 to reference genome mm9. Mapped reads were filtered for mapping quality ≥10, and duplicates were removed.
Reads were extended to a length of 300bp and a scaled (one million / total of unique reads) coverage was computed using genomeCoverageBed from bedtools. We then produced the bigwig files using bedGraphToBigWig tool from UCSC.
Genome_build: mm9
Supplementary_files_format_and_content: bigwig
Submission date Jul 25, 2016
Last update date May 15, 2019
Contact name Guillaume Andrey
Phone +41223795703
Organization name University of Geneva
Department Department of Genetic Medicine and Development
Street address Rue Michel-Servet 1
City Geneva
ZIP/Postal code 1211
Country Switzerland
Platform ID GPL17021
Series (2)
GSE84793 Characterization of hundreds of regulatory landscapes in developing limbs reveals two regimes of chromatin folding [ChIP-seq]
GSE84795 Characterization of hundreds of regulatory landscapes in developing limbs reveals two regimes of chromatin folding
BioSample SAMN05438926
SRA SRX1975284

Supplementary file Size Download File type/resource 66.8 Mb (ftp)(http) BW
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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