 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Dec 01, 2016 |
| Title |
Kdm5A 4 hour exp. 2 |
| Sample type |
SRA |
| |
|
| Source name |
3T3-L1_Kdm5A 4 hour
|
| Organism |
Mus musculus |
| Characteristics |
cell line: 3T3-L1
cell type: Preadipocyte cell line
time point: 4 hours post induction
chip antibody: KDM5A antibody (ab70892; Abcam)
|
| Treatment protocol |
Two days post confluency (day 0), 3T3-L1 cells were induced to differentiate using a standard cocktail of adipogenic inducers (dexamethasone, insulin, 3-isobutyl-1-methylxanthine, and fetal bovine serum). Two days post induction (day 2), medium was changed to DMEM supplemented with fetal bovine serum and insulin. Cells were maintained in DMEM supplemented with fetal bovine serum for the remainder of the differentiation. For knock down experiments, pre-confluent 3T3-L1 cells were transduced 4 days before induction of differentiation by incubation for 24 hours at 37°C with medium containing lentiviral constructs (shKDM5A/B/C or control shLuc) and 6 μg/mL polybrene (Sigma). After 24 hours the medium was changed to normal growth medium.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP experiments were performed in 3T3-L1 cells essentially as previously described (Siersbæk et al. 2012, MCB, 32: 3452-3463). For cross-linking of chromatin, 0.2 mM disuccinimidyl glutarate (DSG) was applied (Proteochem, Denver, CO) for 45 min followed by cross-linking using 1% formaldehyde for 10 min.
ChIP-seq libraries were constructed according to the manufacturer's instructions (Illumina) as described in (Nielsen R, Mandrup S, 2014 Genome-Wide Profiling of Transcription Factor Binding and Epigenetic Marks in Adipocytes by ChIP-seq. Methods in Enzymology 2014, Vol. 537, pp. 261-279).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 1500 |
| |
|
| Description |
processed data file: KDM5A_all_peaks_tags.txt
|
| Data processing |
Sequence reads were aligned to the mouse reference genome (version mm9) with STAR (Dobin et al. 2013, Bioinformatics , 29: 15) set to not map across splice junctions (--alignIntronMax 1 --outSJfilterIntronMaxVsReadN 0)
KDM5A ChIP-seq and matching Input tag counts were normalized to 10M reads using HOMER.
Regions enriched for KDM5A in the individual libraries were called using HOMER with the ‘-style histone’ and '-F 10' settings. For the individual KDM5A region files, overlapping peak regions were merged into one region and KDM5A peaks at each time point were identified as having 10-fold more average normalized tags over two replicates relative to the input control. Promoter proximal regions were identified as being within ±1kb of transcriptional start site (TSS) and distal binding sites between 5 and 50 kb away from TSS.
Tag directories for each ChIP-seq library were generated using HOMER. BedGraph files were created using HOMER (Heinz S, et al. 2010. Mol. Cell 38:576 –589) for visualization of binding profiles
Genome_build: mm9
Supplementary_files_format_and_content: bedGraph formatted file that can then be uploaded as a custom track to the UCSC genome browser
KDM5A_all_peaks_tags.txt: Table containing all identified KDM5A binding sites, including normalized tag counts for KDM5A replicate libraries
KDM5C_all_peaks_tags.txt: Table containing all identified KDM5C binding sites, including normalized tag counts for KDM5C replicate libraries
|
| |
|
| Submission date |
Nov 15, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Ann-Sofie Bøgh Brier |
| E-mail(s) |
a.boegh@me.com
|
| Phone |
+4522969284
|
| Organization name |
University of Southern Denmark, Odense
|
| Department |
Department of Biochemistry and Molecular Biology
|
| Lab |
Mandrup lab
|
| Street address |
Campusvej 55
|
| City |
Odense M |
| ZIP/Postal code |
5230 |
| Country |
Denmark |
| |
|
| Platform ID |
GPL18480 |
| Series (1) |
| GSE84410 |
The KDM5 family is required for activation of pro-proliferative cell cycle genes during adipocyte differentiation |
|
| Relations |
| BioSample |
SAMN06017317 |
| SRA |
SRX2348509 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2391499_SM453_AGTTCC_KDM5A_4h_rep2.TD.bedgraph.gz |
34.4 Mb |
(ftp)(http) |
BEDGRAPH |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
|
|
|
|
 |
