|
Status |
Public on Dec 06, 2016 |
Title |
CTCF-FL-E115-Wt-Mm-Rep1-L8227-1 |
Sample type |
SRA |
|
|
Source name |
Embryonic Limb Bud
|
Organism |
Mus musculus |
Characteristics |
tissue: forelimb developmental stages: E11.5 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.
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|
|
Library strategy |
ChIP-Seq |
Library source |
genomic |
Library selection |
ChIP |
Instrument model |
Illumina HiSeq 2500 |
|
|
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
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Submission date |
Jul 25, 2016 |
Last update date |
May 15, 2019 |
Contact name |
Guillaume Andrey |
E-mail(s) |
guillaume.andrey@unige.ch
|
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 |
|
Relations |
BioSample |
SAMN05438907 |
SRA |
SRX1975279 |