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Sample GSM1967874 Query DataSets for GSM1967874
Status Public on Jun 03, 2016
Title N20 ChIP rep2
Sample type SRA
Source name K562 cells
Organism Homo sapiens
Characteristics passage: 3-5
chip-antibody: N-20 (Santa Cruz)
cell line: K562
Treatment protocol This protocol was adapted from the laboratory of Richard Myers (Johnson et al., Science, 2007). The protocol was followed except for some exceptions. 3x107 K562 cells were crosslinked for 7.5 min with 1% methanol-free formaldehyde (Pierce™ 16% Formaldehyde (w/v), Methanol-free, Thermo Scientific). The chromatin was sonicated on a BioRuptor Next Gen (Diagenode) at high power for 40 cycle of 30’’/30’’ ON/OFF. Antibodies were bound to Dynabeads Protein G (Thermo Scientific) following the manufacturer’s instructions. 1.5 μg of the N-20 antibody (Santa Cruz, sc-899) was bound to 1.5 mg Dynabeads. Samples were immunoprecipitated overnight. Illumina libraries were prepared with NEBNext Ultra DNA Library Prep kit. Samples were sequenced on an Illumina HiSeq 1500 machine.
Growth protocol K562 cells were acquired from DSMZ (Braunschweig, Germany). Cells were grown in RPMI 1640 medium (Gibco) supplemented with 10% heat-inactivated FBS (Gibco) and 1% Penicillin/Streptomycin (100x, PAA) at 37°C under 5% CO2
Extracted molecule genomic DNA
Extraction protocol NEBNext Ultra DNA Library Prep Kit for Illumina
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 1500
Data processing Sequencing data processing. Paired-end 50 base reads with additional 6 base reads of barcodes were obtained for each of the samples, i.e. total RNA (RNA-Seq), total fragmented RNA (RNA-Frag-Seq), labeled RNA (4sU-Seq), labeled fragmented RNA (TT-Seq) and ChIP-Seq samples (N20 and S2P). Reads were demultiplexed and mapped with STAR 2.3.0 (for RNA-Seq) (Dobin and Gingeras, Current protocols in bioinformatics, 2015) and Bowtie 2.1.0. (Langmead and Salzberg, Nat Methods, 2012) (for ChIP-Seq) to the hg20/hg38 (GRCh38) genome assembly (Human Genome Reference Consortium). Samtools (Li et al., Bioinformatics, 2009) was used to quality filter SAM files, whereby alignments with MAPQ smaller than 7 (-q 7) were skipped and only proper pairs (-f99, -f147, -f83, -f 163) were selected. Further processing of the RNA-Seq data was carried out using the R/Bioconductor environment. For visualization purposes piled-up read counts for every genomic position were summed up over replicates, size factors for each condition were calculated as described in (Anders and Huber, Genome Biol, 2010) and used to correct for library size and sequencing depth variations.
Genome_build: hg20/hg38 (GRCh38)
Supplementary_files_format_and_content: The file binned_count_table.txt gives the fragment counts for all experiments using the binning_anno.txt annotation file
Supplementary_files_format_and_content: The Series file README.txt includes information about the contents of the processed data files.
Submission date Dec 08, 2015
Last update date May 15, 2019
Contact name Björn Schwalb
Organization name MPI
Street address Fassberg 11
City Göttingen
ZIP/Postal code 37077
Country Germany
Platform ID GPL18460
Series (1)
GSE75792 TT-Seq captures the human transient transcriptome
BioSample SAMN04325074
SRA SRX1470573

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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