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| Status |
Public on Mar 07, 2019 |
| Title |
WT_NSCs_H3K4me1_ChIPSeq |
| Sample type |
SRA |
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| Source name |
wild type NSC
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| Organism |
Mus musculus |
| Characteristics |
strain background: mixed C57BL/6 x DBA
cell type: forebrain-derived neural stem cells (NSCs)
genotype/variation: wild type
passages: passage 3 to 7
chip antibodies: Ab8894 (Abcam) for H3K4me1.
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| Growth protocol |
P0 brain-derived NSC cultures were obtained from dissected telencephalon of six wild type and six Sox2-deleted mice, and grown, as described in (Favaro et al., 2009) and (Zhang et al., 2013). Cells were grown in parallel for two passages in 10 ng/ml Epidermal Growth Factor (EGF) and 10 ng/ml basic Fibroblast Growth Factor (bFGF), followed by cell dissociation and expansion in the presence of EGF, but not bFGF, for 3-4 more passages, as described in Zhang et al. (2013), for optimal maintenance of mutant NSC (Favaro et al., 2009).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For each line, neurospheres were dissociated and 4 million single cells were crosslinked with 1% formaldehyde for 10 minutes at room temperature. Reaction was quenched with 0.125 M Glycine, cells were washed with cold PBS and lysed according to Vermunt et al., 2016. Nuclei pellets were resuspended in 160μl sonication buffer and divided over two microtubes for shearing in the Covaris S series with the following settings for 12 cycles of 60 seconds: intensity 3, duty cycle 20%, 200 cycles/bursts. Chromatin immunoprecipitation steps after sonication were performed as described previously (Vermunt et al., 2016) using 50 μl DynaI protein G beads that were preincubated with 5 μg Ab4729 (Abcam) for H3K27ac or 5 μg Ab8894 (Abcam) for H3K4me1. Whole cell extract of 4 million cells was split onto both antibodies, resulting in the use of 2 million cells per ChIP.
Libraries were made using the Illumina Truseq DNA library protocol and sequencing was done at the MIT BioMicro Center (http://openwetware.org/wiki/BioMicroCenter).
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
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| Data processing |
Basecalls performed using CASAVA
ChIP-seq reads were aligned to the mm9 genome assembly using Bowtie version 0.12.5 with at most one substitution. Reads mapping at multiple positions were discarded
MACS2 was used for peak calling (p-value threshold = 10-5, extsize = 400, local lambda = 100,000) and narrowpeaks were extended to a minimum of 2000 basepairs (bps) to match peak resolution. Overlapping enriched regions were merged.
Genome_build: mm9
Supplementary_files_format_and_content: NR_H3K27ac_EnrichedRegions_Sort.bed and NR_H3K4me1_EnrichedRegions_Sort.bed. Enriched regions for each histone modification in wild type NSC cells
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| Submission date |
Nov 26, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Federico Zambelli |
| Organization name |
University of Milan
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| Department |
Department of Biosciences
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| Street address |
Via Giovanni Celoria 26
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| City |
Milano |
| ZIP/Postal code |
20133 |
| Country |
Italy |
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| Platform ID |
GPL9250 |
| Series (2) |
| GSE90558 |
Genome-wide maps of chromatin state in neural stem/progenitor cells (NSC) from mouse neonatal forebrain [ChIP-seq] |
| GSE90561 |
Sox2 is required for functional chromatin connectivity in brain-derived neural stem cells. |
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| Relations |
| BioSample |
SAMN06060054 |
| SRA |
SRX2378794 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2406793_NR_H3K4me1_EnrichedRegions_Sort.bed.gz |
930.2 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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