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Sample GSM335214 Query DataSets for GSM335214
Status Public on Jun 01, 2009
Title H3K9ac_in4hPI_H3
Sample type genomic
 
Channel 1
Source name H3K9ac ChIP DNA from EL4 cells stimulated for 4h with PMA/IoM
Organism Mus musculus
Characteristics Antibody: H3K9ac
Cell Line: EL4
Treatment: PMA (10 ng/ml phorbol myristate acetate) and Ionomcyin (1 microM)
Time: 4h
Treatment protocol EL4 cells were stimulated with PMA (10 ng/ml phorbol myristate acetate) and Ionomcyin (1 microM) for 0h, 0.5h or 4h
Growth protocol EL-4 were cultured in RPMI 1640 medium with 10 mM HEPES, 10% fetal calf serum (FCS; CSL, Australia), 120 microg/ml penicillin, and 16 microg/ml gentamycin.
Extracted molecule genomic DNA
Extraction protocol ChIP assays were performed as previously described (Chen et al. Mol. Cell. Biol (2005) 25:3209) with some modifications. Briefly, cells were harvested and cross-linked with formaldehyde. Cells were lysed and then sonicated using the Bioruptor (Diagenode) to give fragments between 200bp to 1000bp lengths. Samples were precleared with protein-A agarose/salmon sperm DNA beads (Upstate) then immunoprecipitated with 2.5 microg anti-histone-H3 (Abcam), 4 microg anti-acetyl-H3K9 (Upstate) or mock immunoprecipiatated. Immune complexes were recovered using salmon sperm DNA/protein A-agarose, washed and eluted. Following cross-link reversal and proteinase K treatment immunoprecipitated DNA was purified by phenol–chloroform extraction and ethanol precipitation.10ng ChIP DNA was amplified with the Whole Genome Amplification (WGA) kit from Sigma as per manufacturers instructions but with incorporation of dUTP.
Label biotin
Label protocol Samples (7.5μg of DNA) were enzymatically fragmented and labeled using the WT Labeling Kit (Affymetrix) to add biotinylated compound to the ends of the fragments
 
Channel 2
Source name Pan Histone H3 ChIP DNA from EL4 cells stimulated for 4h with PMA/IoM
Organism Mus musculus
Characteristics Antibody: Histone H3
Cell Line: EL4
Treatment: PMA (10 ng/ml phorbol myristate acetate) and Ionomcyin (1 microM)
Treatment protocol EL4 cells were stimulated with PMA (10 ng/ml phorbol myristate acetate) and Ionomcyin (1 microM) for 0h, 0.5h or 4h
Growth protocol EL-4 were cultured in RPMI 1640 medium with 10 mM HEPES, 10% fetal calf serum (FCS; CSL, Australia), 120 microg/ml penicillin, and 16 microg/ml gentamycin.
Extracted molecule genomic DNA
Extraction protocol ChIP assays were performed as previously described (Chen et al. Mol. Cell. Biol (2005) 25:3209) with some modifications. Briefly, cells were harvested and cross-linked with formaldehyde. Cells were lysed and then sonicated using the Bioruptor (Diagenode) to give fragments between 200bp to 1000bp lengths. Samples were precleared with protein-A agarose/salmon sperm DNA beads (Upstate) then immunoprecipitated with 2.5 microg anti-histone-H3 (Abcam), 4 microg anti-acetyl-H3K9 (Upstate) or mock immunoprecipiatated. Immune complexes were recovered using salmon sperm DNA/protein A-agarose, washed and eluted. Following cross-link reversal and proteinase K treatment immunoprecipitated DNA was purified by phenol–chloroform extraction and ethanol precipitation.10ng ChIP DNA was amplified with the Whole Genome Amplification (WGA) kit from Sigma as per manufacturers instructions but with incorporation of dUTP.
Label biotin
Label protocol Samples (7.5μg of DNA) were enzymatically fragmented and labeled using the WT Labeling Kit (Affymetrix) to add biotinylated compound to the ends of the fragments
 
 
Hybridization protocol Approximately 7.5μg of DNA was hybridzed per array using the Affymetrix hybridization kit. Arrays were hybridized for 16 hours at 45° at 60rpm using an Affymetrix hybridization oven and washed with Fluidics station 450 protocol FS450_0001
Scan protocol Arrays were scanned on an Affymetrix Scanner 3000 7G
Description H3K4ac ChIP in EL4 cells stimulated with PMA and Ionomcyin for 4 hours, using using total histone 3 ChIP as control, 2 biological replicates
Data processing All analysis was performed with NCBI build 36 of the mouse genome. The model-based analysis of tiling array (MAT) algorithm (Johnson WE et al PNAS 2006) was used to find regions of H3K9Ac with a bandwidth of 250bp and a max gap of 150bp. Either the matching total input DNA or H3 chips (as stated) were used as control samples.
 
Submission date Oct 20, 2008
Last update date Jan 04, 2012
Contact name Kristine Hardy
E-mail(s) kristine.hardy@anu.edu.au
Organization name University of Canberra
Lab Cytokine Gene Expression
Street address University of Canberra
City Bruce
State/province ACT
ZIP/Postal code 0200
Country Australia
 
Platform ID GPL5811
Series (2)
GSE13277 ChIP-on-chip examination of inducible genes in T cells
GSE13279 Defining the chromatin signature of inducible genes in T cells

Supplementary file Size Download File type/resource
GSM335214_H3_4h_rep1.CEL 44.8 Mb (ftp)(http) CEL
GSM335214_H3_4h_rep3.CEL 44.8 Mb (ftp)(http) CEL
GSM335214_H3k9Ac_h3c_4_2reps.Mm_PromPR_v02-1_NCBIv36.NR.bpmap_matscore.bar 32.4 Mb (ftp)(http) BAR
GSM335214_H3k9Ac_h3c_4_2reps.bed 4.1 Mb (ftp)(http) BED
GSM335214_K9_4h_rep1.CEL 44.8 Mb (ftp)(http) CEL
GSM335214_K9_4h_rep3.CEL 44.8 Mb (ftp)(http) CEL
Processed data provided as supplementary file

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