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Sample GSM1443813 Query DataSets for GSM1443813
Status Public on Dec 12, 2014
Title KLF11 hMADS white rep1, day 19
Sample type SRA
 
Source name human adipose-derived stem cells
Organism Homo sapiens
Characteristics cell line: hMADS-3
cell type: human adipose-derived stem cells
chip antibody: anti-KLF11 (10D8; Novus Biologicals)
Treatment protocol Two days post confluence (day 0) hMADS cells were induced to differentiate in DMEM/Ham`s F12 (Lonza) supplemented with 10 μg/ml transferrin, 1 μM dexamethasone, 500 μM 3-isobutyl-1-methylxanthine (MIX), 0.85 μM insulin, and 0.2 nM T3. At day 3 of differentiation, MIX and dexamethasone were omitted from the media and 0.5 μM rosiglitazone was added until day 9 of differentiation and again from day 13-16 for induction of brite adipocytes. Control white adipocytes were treated with DMSO from day 13-16. From day 16 and forth the differentiation medium of both white and brite adipocytes was depleted of rosiglitazone. White and brite hMADS adipocytes were harvested on day 19, unless stated otherwise.
Growth protocol Human multipotent adipose-derived stem (hMADS-3) cells were cultured in DMEM (Lonza, low glucose) supplemented with 10% fetal bovine serum (Lonza), 10 mM Hepes, 2 mM L-glutamine (Lonza), penicillin (62.5 μg/ml), streptomycin (100 μg/ml), and 2.5 ng/ml hFGF2 (Peprotech).
Extracted molecule genomic DNA
Extraction protocol ChIP experiments were performed in mature hMADS adipocytes 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
 
Data processing Alignment:
Sequence reads were collapsed using FASTX-Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/). Bowtie (Langmead B, et al., Genome Biol. 10:R25) was used to align each collapsed library to the human reference genome (version hg19) with the following parameters: “-m 3 –best –strata”, with all other parameters default.
Peak finding and visualization:
Regions enriched for KLF11 signal in white and brite hMADS adipocytes were identified using HOMER (Heinz S, et al. 2010. Mol. Cell 38:576–589) with the “-factor” and “-size given” settings and a 0.1% FDR threshold. An additional stringency filter was applied, so only peaks >5 fold over the matching input control were kept for further analyses. Merged peak files were generated from the individual KLF11 peak files if the center of two or more peaks were within 250 bp. Peaks from these merged peak files were only included if having more than 15 tags per 10 M tags in a 400 bp window around the center of each merged peak. BedGraph files were created using HOMER for visualization of binding profiles.
Genome_build: hg19
 
Submission date Jul 23, 2014
Last update date May 15, 2019
Contact name Anne Loft
E-mail(s) anlo@bmb.sdu.dk
Organization name University of Southern Denmark
Department Biochemistry & Molecular Biology
Street address Campusvej 55
City Odense M
ZIP/Postal code 5230
Country Denmark
 
Platform ID GPL18460
Series (1)
GSE59703 Browning of human adipocytes requires KLF11 and reprogramming of PPARγ super-enhancers
Relations
BioSample SAMN02934493
SRA SRX660087

Supplementary file Size Download File type/resource
GSM1443813_SM640.KLF11W.rep1.bedGraph.gz 27.2 Mb (ftp)(http) BEDGRAPH
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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