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Status |
Public on Nov 17, 2014 |
Title |
H3K4me3 ChIP-seq in Suz12GT (aka Suz12(Bgal/Bgal)) ESCs |
Sample type |
SRA |
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Source name |
ES cells
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Organism |
Mus musculus |
Characteristics |
cell line: Suz12GT time: d0 cell type: ESCs genotype/variation: mutant in Suz12 mutant cell line reference: Pasini et al., 2007 strain: 129/Ola chip antibody: anti-H3K4me3 (Millipore, catalog# 07-473)
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Treatment protocol |
N/A
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Growth protocol |
ESCs were plated with irradiated murine embryonic fibroblasts (MEFs) and grown under typical ESC conditions on gelatinized tissue culture plates. Briefly, cells were grown in Knockout DMEM supplemented with 15% fetal bovine serum, leukemia inhibitory factor (LIF), non-essential amino acids, L-glutamine, and penicillin/streptomycin as previously described (Boyer et al., 2006). Cells were then preplated - plated on cell culture plates without gelatin for 20 minutes to allow the MEFs to attach and thus deplete the cell population of MEFs - before collection for ChIP.
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Extracted molecule |
genomic DNA |
Extraction protocol |
The ChIP protocol has been adapted from previous studies (Lee et al., 2006; Boyer et al., 2006, Creyghton et al., 2008) with the following modification: Diagenode Bioruptor was used to sonicate with 30 cycles of 30 sec on (high), 30 sec off. The samples were sonicated in 15ml polystyrene tubes at 4°C while samples were immersed in ice cold water. Millipore 07-473 antibody was used in the ChIP. The ChIP DNA was then used to prepare sequencing libraries as described in Schmidt et al., 2009. For this study, a 1:40 dilution of Single End read adapters (Illumina) was used in the ligation reaction. Each sample was amplified for 18 cycles. The samples were then examined for proper size and structure by Bioanalyzer (Agilent) and qPCR.
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Library strategy |
ChIP-Seq |
Library source |
genomic |
Library selection |
ChIP |
Instrument model |
Illumina Genome Analyzer IIx |
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Description |
Suz12GT_d0_H3K4me3
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Data processing |
Illumina Offline BaseCaller1.9.3 software used for basecalling. ChIP-seq reads were aligned to the mm9 genome assembly using Bowtie 0.12.3, with the following parameters --solexa1.3-quals --best --strata -m 1 -n 2 -p 8 -S. Sequences were extended +200 bp for histone marks and allocated in 25-bp bins. A Poissonian model was used to determine statistically enriched bins with a P-value threshold set at 1x10-9 as described previously (Marson et al., 2008). Additionally, we required that genomic bins were at least 5 fold over input to be considered enriched peaks. Genome_build: mm9 Supplementary_files_format_and_content: wig files were generated using an in-house perl script interacting with Bedtools (as described in Wamstad et al. Cell 2012).
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Submission date |
Sep 09, 2014 |
Last update date |
May 15, 2019 |
Contact name |
Laurie A Boyer |
E-mail(s) |
lboyer@mit.edu
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Phone |
617 324-3335
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Organization name |
Massachusetts Institute of Technology
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Department |
Biology
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Lab |
Boyer
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Street address |
77 Massachusetts Avenue
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City |
Cambridge |
State/province |
MA |
ZIP/Postal code |
02139 |
Country |
USA |
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Platform ID |
GPL11002 |
Series (2) |
GSE53508 |
PRC2 coordinates lineage fidelity and DNA methylation during ESC differentiation |
GSE61248 |
Change in H3K4me3 in Suz12(Bgal/Bgal) ESCs, and over Spinal Motor Neuron differentiation (ChIP-Seq) |
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Relations |
SRA |
SRX697787 |
BioSample |
SAMN03031396 |
Supplementary file |
Size |
Download |
File type/resource |
GSM1500754_ST1309_Suz12GTd0_K4.WIG.gz |
6.2 Mb |
(ftp)(http) |
WIG |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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