|
| Status |
Public on Sep 20, 2013 |
| Title |
H3K27Ac_ChIPSeq_rep2_B |
| Sample type |
SRA |
| |
|
| Source name |
Peripheral blood cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: CD14+ monocytes
chip antibody: H3K27ac (Abcam ab-4729)
experimental condition: Cultured with MCSF (10ng/ml) and IFN-gamma (100U/ml) for 24 hours.
|
| Treatment protocol |
Cells were treated with or without IFN-g (100U/ml) for 24 hours, and then stimulated with LPS (50ng/ml) for 3 or 6 hours as indicated.
|
| Growth protocol |
Peripheral blood mononuclear cells were obtained from the blood of healthy donors by density gradient centrifugation using Ficoll (Invitrogen) and a protocol approved by the Hospital for Special Surgery Institutional Review Board. CD14+ human monocytes were purified from PBMCs by positive selection using anti-CD14 magnetic beads as recommended by the manufacturer (Miltenyi Biotec). Monocytes were cultured in Roswell Park Memorial Institute (RPMI) 1640 medium (Invitrogen) supplemented with 10% heat-inactivated defined FBS (HyClone), penicillin/streptomycin (Invitrogen), L-glutamine (Invitrogen), and 10 ng/mL human macrophage colony-stimulating factor (M-CSF; Peprotech).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Sonicated cell lysates of nuclei and histone-DNA complexes were isolated with antibody.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, Klenow polymerase and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (3’ to 5’ exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was purified using MinElute PCR Purification kit and PCR amplified with Illumina primers for 15 cycles. Library fragments of 200-300 bp were band isolated from an 1.5% agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on an Illumina HiSeq 2000 Sequencer following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
Basecalls performed using CASAVA version 1.7
ChIP-seq reads were aligned to the hg19 genome assembly using BOWTIE with default configurations.
peaks were called using ChIPSeeqer with the following setting: STAT1 -t_15 -t_fold 2; H3K27Ac -t_20 -t_fold 3 ; IRF1 -t_15 -t_fold 2;
Genome_build: hg19
Supplementary_files_format_and_content: wig files were generated using ChIPSeeqer
The headers for peak *txt files are as follows (from the left):
Chromosome : chromosome name
Start_Position : the first (genomic) coordinate of the peak
End_Position : the second (genomic) coordinate of the peak
Avg_p-value : average log p-value of the nucleotides in the normalized peak region
Score : score estimated as the average ChIP reads/length of peak (minus the average INPUT reads/length of peak - if INPUT is available)
Posmaxpeakheight : the position of the maximum height of the peak
Maxpeakheight : the maximum peak height (in reads)
RelPosMaxPeakHeight(%) : the relative position of the maximum height of the peak, e.g., 50% means the highest point is at the middle of the peak
Peak_Size : the size of the peak (in bp)
Mid_point : the middle position of the peak
Summit_dist_from_mid : the distance of the maximum height from the middle of the peak
|
| |
|
| Submission date |
Dec 21, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Lionel Ivashkiv |
| Organization name |
Hospital for Special Surgery
|
| Street address |
535 East 70th Street
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10021 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE43036 |
Synergistic Activation of Inflammatory Cytokine Genes by Priming of Regulatory DNA Elements for Increased Transcription in Response to TLR Signaling |
|
| Relations |
| SRA |
SRX212658 |
| BioSample |
SAMN01874817 |
