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Sample GSM1625985 Query DataSets for GSM1625985
Status Public on Feb 28, 2016
Title Input - baseline Macrophages
Sample type SRA
Source name primary human macrophages
Organism Homo sapiens
Characteristics cell type: peripheral blood derived macrophage
treatment: GM-CSF (72h)
chip antibody: rabbit polyclonal anti-H3K4me1 antibody (Abcam cat.# 8895)
Treatment protocol Fractions of baseline macrophages were further stimulated for 72h in culture medium with the addition of three stimulatory cocktails containing IFN-γ (200U/ml), IL-4 (500U/ml) or a combination of TNF-α (800U/ml), PGE2 (1µg/ml) and Pam3CSK4 (1µg/ml) to differentiate macrophages into subtypes
Growth protocol MACS beads isolated CD14 primary human monocytes were cultured for 72 h with granulocyte/macrophage colony stimulating factor (GM-CSF; 500U/ml) in RPMI1640 medium containing 10 % fetal calf serum (FCS) and 1% Penicillin/Streptomycin to generate unstimulated baseline macrophages
Extracted molecule genomic DNA
Extraction protocol Chromatin from 0.5x10E6 macrophages was cross-linked for ChIP reactions with 1 % formaldehyde. Nuclei were lysed and chromatin was sheared with the Covaris S220 ultrasound system. For ChIP antibody reactions polyclonal rabbit antibodies were used (H3K4me1 (Abcam), H3K27Ac (Abcam), H3K27me3 (Millipore)). Washing and DNA purifications as well as size selections were performed with AMPure XP SPRI beads (Beckman Coulter).
Published library construction protocol was adapted (Blecher-Gonen et al., Nature Protocols, 2013) and performed with 0.5 ng ChIP DNA. Barcodes from the NEXTflex adapter oligonucleotides kit (Bioo Scientific) were ligated to the ChIP-seq DNA fragments. Final DNA purification with additional size selection was performed with SPRI beads. ChIP-seq libraries were sequenced on the HiScanSQ/Hiseq 1000 sequencer (Illumina)
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 1500
Data processing Basecalls performed using CASAVA software version 1.8
ChIP-seq reads were aligned to the NCBI Build GRCh36/UCSC Build hg18 genome assembly using Bowtie v0.12.8 with following options: -t -q -e 70 -l 28 -n 2 --best --maxbts 125 -S
After ChIP-seq quality control using HOMER v4.3, reads of biological replicates were pooled, and peaks were called using HOMER 4.3 with script with pooled Input samples used as background control files using default options and following additional settings: -style histone -size 1000 -minDist 2500 -nfr
Annotation of peak positions and counting of normalized tag counts in called regions was performed with HOMER v4.3 script.
Genome_build: hg18
Supplementary_files_format_and_content: Annotated HOMER called peak positions in tab-delimited text format
Submission date Mar 05, 2015
Last update date May 15, 2019
Contact name Joachim Schultze
Organization name LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)
Department Genomics and Immunoregulation
Street address Carl-Troll-Strasse 31
City Bonn
State/province NRW
ZIP/Postal code 53115
Country Germany
Platform ID GPL18460
Series (2)
GSE66594 Epigenomic landscapes of human inflammation associated macrophages [ChIP-seq]
GSE66595 Epigenomic landscapes of human inflammation associated macrophages
BioSample SAMN03389893
SRA SRX903258

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data not provided for this record

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