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Sample GSM2538831

Query DataSets for GSM2538831

Status

Public on Apr 25, 2018

Title

H3K9me3_ChIPSeq [C3649]

Sample type

SRA

Source name

induced pluripotent stem cells, H3K9me3 ChIP

Organism

Pan troglodytes

Characteristics

cell line: C3649
cell type: induced pluripotent stem cells
Sex: Male
population: Chimpanzee
chip antibody: H3K9me3 (Abcam, 8898, GR131093-3)

Growth protocol

Human and chimpanzee feeder-independent induced pluripotent stem cell lines were maintained at 70% confluence on Matrigel hESC-qualified Matrix (354277, Corning, Bedford, MA, USA) at a 1:00 dilution. Cells were cultured in Essential 8 Medium (A1517001, ThermoFisher Scientific, Waltham, MA, USA) at 37°C with 5% (vol/vol) CO2 with daily media changes. Cells were passaged by enzyme-free dissociation (0.5 mM EDTA, 300 mM NaCl in PBS), and seeded with ROCK inhibitor Y-27632 (ab120129, Abcam, Cambridge, MA, USA).

Extracted molecule

genomic DNA

Extraction protocol

ChIP-seq experiments were largely performed according to a published protocol (Schmidt et al. 2009). Briefly, 30 million cells were cross-linked with 1% formaldehyde for 10 min at room temperature followed by quenching with 2.5 M glycine. Following cell lysis and sonication (Covaris S2: 4 min, duty cycle 10%, 5 intensity, 200 cycles per burst in 4x 6 x 16 mm tubes per individual), lysates were incubated with 5 μg H3K9me3 antibody (8898, Abcam) overnight.
Libraries were prepared using the Illumina TruSeq ChIP Sample Preparation kit. Libraries were sequenced on the HiSeq2500 following the manufacturer's protocols.

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

Illumina HiSeq 2500

Description

Processed data file: H3K9me3_counts_hg19.txt
Processed data file: H3K9me3_counts_panTro3.txt

Data processing

Paired-end ChIP-seq reads from each species were mapped to their respective genome (hg19 or panTro3) using BWA (version 0.7.12) (Li and Durbin 2009) and a mapping quality filter of MAQ >10. PCR duplicates were removed by samtools (version 0.1.19)(Li et al. 2009). Properly-paired reads only were selected for further analysis.
ChIP and Input reads from each individual were used to call peaks using MACS2 (Zhang et al. 2008) with the broadpeak option at various qvalue cut-offs. A lenient threshold of qvalue=0.1 was used in subsequent analysis to maximize the number of regions in each species. Peaks called in each species were concatenated to generate a final peak set across both species.
Peak coordinates in each species were then converted to the alternative genome using liftOver (Speir et al. 2016) and the best reciprocal chain (http://hgdownload.cse.ucsc.edu/goldenPath/hg19/vsPanTro3/reciprocalBest/) between hg19 and panTro3. Peak regions that could be reciprocally and uniquely mapped between the two genomes were kept to generate a list of orthologous peaks.
The number of reads falling into these orthologous peak regions was determined using HTSeq (Anders et al. 2015).
Genome_build: hg19 (GRCh37), panTro3 (Pan_troglodytes-2.1.3)
Supplementary_files_format_and_content: *txt: H3K9me3 ChIP-seq read counts for all orthologous regions in all samples with region coordinates on both the human genome (hg19) and the chimpanzee genome (panTro3).

Submission date

Mar 16, 2017

Last update date

May 15, 2019

Contact name

Michelle C Ward

Organization name

University of Chicago

Department

Human Genetics

Lab

Gilad