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| Status |
Public on Sep 04, 2015 |
| Title |
H2BGlcNAc_Day3 |
| Sample type |
SRA |
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| Source name |
Pre-adipocytes from liposuction material, cultured, 3 days after adipogenic stimulation (Day 3).
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| Organism |
Homo sapiens |
| Characteristics |
passages: Cells in passage 5-7
initial cell type: pre-adipocytes
cell differentiation status: Non-proliferating, differentiated for 3 days
chip antibody: anti-H2B (glcnac S112) Abcam ab130951
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| Growth protocol |
Primary ASCs isolated from liposuction material were cultured in T175 flasks under proliferative conditions in high-glucose DMEM (4.5 g/l; Life Technologies-BRL) containing 20% FBS and 2 ng/ml basic fibroblast growth factor (Sigma-Aldrich). Cells at passage 5-7 were used for adipogenic induction. When cells reached confluency, adipogenic differentiation was induced by replacing growth medium with DMEM/F12 containing 10% FBS and by addition of 0.5 µM 1-methyl-3 isobutylxanthine (Dumex Alpharma), 1 µM dexamethasone (Dumex Alpharma), 10 µg/ml insulin (Novo Nordisk) and 100 µM (Dumex Alpharma) indomethacin for up to 9 days.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For ChIP-seq: Cells (10e7 cells/ChIP) were harvested and fixed with 1% formaldehyde for 10 min before quenching with 125 mM glycine. Cells were lysed for 30 min at 4C on a rotator in RIPA buffer (140 mM NaCl, 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X-100, 0.1% SDS, 0.1% deoxycholate, 1 mM PMSF, protease inhibitor cocktail) adjusted to 1% SDS, and sonicated 3 times 15 min in a Bioruptor® (Diagenode). After sedimentation at 10,000 g for 10 min, the supernatant was collected and diluted 10x in RIPA buffer without SDS to constitute input chromatin. Chromatin was incubated overnight at 4oC on a rotator with antibodies coupled to magnetic Dynabeads Protein G. ChIP samples were washed 3 times in ice-cold RIPA buffer. Crosslinks were reversed and DNA eluted for 6 h at 37C in 50 mM NaCl, 20 mM Tris-HCl pH 7.5, 5 mM EDTA, 1% SDS, 0.5 µg/ml RNase A, and 2 µg/ml Proteinase K added twice. DNA was purified and dissolved in H2O.
Sequencing library was prepared according to the Illumina protocol, and sequenced on an Illumina HiSeq2500 at the Norwegian Sequencing Center.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Reads aligned to reference using bowtie version 1.0.0 using parameters –best
bigWig files were generated by getting a ratio of IP over Input in 1 kb bins
Peaks called using Enriched Domain Detector
Genome_build: hg19
Supplementary_files_format_and_content: bigWig files for viewing in Genome Browser; bed files with four columns: chromosome, start position, end position and score; csv file created from cuffdiff output file genes.fpkm_tracking
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| Submission date |
Nov 17, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Philippe Collas |
| Organization name |
University of Oslo
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| Department |
Institute of Basic Medical Sciences
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| Street address |
PO Box 1112 Blindern
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| City |
Oslo |
| ZIP/Postal code |
0317 |
| Country |
Norway |
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| Platform ID |
GPL16791 |
| Series (1) |
| GSE63346 |
Pre-patterning of differentiation-driven nuclear lamin A/C-associated chromatin domains by GlcNAcylated histone H2B |
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| Relations |
| BioSample |
SAMN03196930 |
| SRA |
SRX760965 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1546364_H2BGlcNAc_D3_peaks.bed.gz |
9.8 Kb |
(ftp)(http) |
BED |
| GSM1546364_H2BGlcNAc_D3_ratios.bw |
26.4 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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