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Sample GSM1175115 Query DataSets for GSM1175115
Status Public on Aug 13, 2013
Title H3K27me3_Basal
Sample type SRA
Source name Neural progenitor cells
Organism Mus musculus
Characteristics strain: FVB/N
cell type: Primary adult neural progenitor cell
passages: 4-9
chip antibody: H3K27me3: Millipore 07-449 lot# DAM1588246
Treatment protocol To induce FOXO3 nuclear accumulation, cells were incubated in low growth factor signaling conditions: Neurobasal A medium supplemented with penicillin/streptomycin/glutamine and 2% B27 for 4 hours, followed by a 1.5 hour incubation with 20 mM LY294002 (LY, Calbiochem-Novabiochem Corp.), a specific PI3K inhibitor, to inhibit residual growth factor signaling.
Growth protocol Adult (12-week-old) mouse NPCs were isolated as previously described (Renault et al., Cell Stem Cell 2009). Briefly, whole mouse forebrains were homogenized and incubated for 30 min in HBSS (Invitrogen) with 1 U/ml DispaseII (Roche), 250 U/ml DNaseI (Sigma) and 2.5 U/ml Papain (Worthington) at 37oC. After mechanical dissociation, cells were purified by sequential 25% and 65% Percoll (Amersham) gradients. Cells were cultured in high growth factor signaling conditions: Neurobasal A (Invitrogen) medium supplemented with penicillin/streptomycin/glutamine (Invitrogen), 2% B27 (Invitrogen) and 20 ng/ml each of FGF2 (Peprotec) and EGF (Peprotec).
Extracted molecule genomic DNA
Extraction protocol Chromatin was cross-linked with 1% formaldehyde for 10 min, followed by quenching with 0.125 M glycine for 5 min. Cells were washed and incubated in swelling buffer (10 mM HEPES pH 7.8, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, 1 mM DTT, 0.5 mM PMSF) for 15 min on ice, followed by nuclei isolation by dounce homogenization. Nuclei were pelleted and resuspended in RIPA buffer (1% NP-40, 0.5% sodium deoxycholate, 1% SDS in PBS pH 7.4), and chromatin was sheared with a Vibra-Cell Sonicator VC130 (Sonics) six times for 30 sec at 60% amplitude.
Single end libraries were generated according to the manufacturer’s instructions (Illumina).
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina Genome Analyzer IIx
Data processing Reads with low quality scores (sequences that have phred <15 on 85% seq) were filtered using the fastx toolbox (fastq_quality_filter -v -Q 33 -q 15 -p 85)
Reads were aligned to mm9 using bowtie-0.12.7 (bowtie -p 4 -S -q -v 2 -m 3 --best --strata)
For chromatin marks, peaks were generated using Macs (version 2) software with "broad option" and with default settings (macs2 --broad)
For transcription factor, peaks were generated using QuEST 2.1 using the highest stringency peak calling parameters (Fold enrichment > 50, bandwidth=30)
Genome_build: mm9
Supplementary_files_format_and_content: H3K27me3, H3K4me1 and H3K4me1 peak files were generated using MACS 2, Ascl1 and FoxO3 peak bed files were generated using QuEST 2.1
Submission date Jun 26, 2013
Last update date May 15, 2019
Contact name Anne Brunet
Organization name Stanford University
Department Genetics
Street address 300 Pasteur Drive,
City Stanford
State/province California
ZIP/Postal code 94305
Country USA
Platform ID GPL11002
Series (1)
GSE48336 FOXO3 shares common targets with ASCL1 genome-wide and inhibits ASCL1-dependent neurogenesis
BioSample SAMN02214079
SRA SRX315276

Supplementary file Size Download File type/resource
GSM1175115_H3K27me3_Minus_pooled_filtered_broad_peaks.bed.gz 518.3 Kb (ftp)(http) BED
SRA Run SelectorHelp
Processed data provided as supplementary file
Raw data are available in SRA

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