{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Sunil Kumar Raghav"],"organism":["Mus musculus"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-10992"],"description":["H3K27ac ChIP-seq was performed to identify active enhancers in different TLR activated control CD8α+ DCs as well as to understand the role of NCoR1 in regulating TLR specific enhacner activity using H3K27ac ChIP-seq performed in NCoR1 KD DCs. MutuDC1940 cell line were treated with TLR3 ligand poly I:C, TLR9 ligand CpG-B and combined stimulation of TLR9 , TLR3, and IFNg for 2hr and 6hr. Recombinant Anti-Histone H3 (acetyl K27) antibody was used to pull down chromatin regions having active enhacner mark in each unstimulated and stimulated condition."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Nucleic Acid Extraction - The ChIP experiments were performed as per the Mayer’s Lab Protocol. The cells were cross-linked using 1% formaldehyde (Sigma) for 10 min at room temperature followed by quenching the reaction using 2.5 M glycine (Sigma) for 10 min. The cells were lysed in the FARHAM lysis buffer and centrifuged at 2000rpm at 4°C for 8 min. The chromatin was fragmented using a Bioruptor (Diagenode) sonicator for 30 min using high amplitude and 30s ON & 30s OFF cycles to obtain 200-500 bp size fragments. The concentration of the chromatin was estimated using a NanoDrop (Thermo) and the chromatin was diluted with a RIPA buffer prepared without protease inhibitor to make 125μg/ml of chromatin for each IP. 30ul of Dyna Magnetic beads (Anti-rabbit) were taken and added to 1ml tube for each IP. 3ul of rabbit monoclonal anti-H3K27ac antibody (Abcam, cat no: ab-177178), were added and incubated at 4°C overnight on a rocker shaker. Next day, the beads were washed six to seven times with LiCl buffer (1% NP-40, 100mM Tris HCl (pH 7.5), 500mM LiCl, 1% Sodium Deoxycholate) followed by two washes with TE buffer (10nM Tris HCl (pH 7.5), 0.1mM EDTA (pH 8)). After removing the wash buffer completely, protein-bound chromatin complexes were eluted from beads for 30 min using 200ul of elution buffers. The eluted chromatin was reverse-crosslinked by overnight incubation on the shaker using 8ul of 5M NaCl. Next day DNA was purified from the reverse cross-linked chromatin by proteinase-K and RNase digestion followed by purification using PCR purification kit (Qiagen).","Sequencing - H3K27ac ChIP sample library were sequenced in Illumina NextSeq-550.","Library Construction - H3K27ac ChIP sample library preparation was performed using an NEB ChIP library preparation Kit","Sample Collection - The CD8α+ lines (mutuDC) were cultured and stable Empty (Control) and NCoR1 shRNA vector (NCoR1 KD) transduced cells were generated as described in previous studies (Fuertes Marraco et al., 2012; Pigni et al., 2018). Two  independent biological replicate of Control and NCoR1 KD cells were treated with CpG-B, poly(I:C) (pIC) and combined CpG+pIC for 2hr and 6hr. Cells were further taken for H3K27ac ChIP-seq."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Sequence Alignment - RAW single end reads were processed for quality check using the FASTQC tool and aligned to the mouse reference genome (mm10) using bowtie2 (2.3.4.2). Duplicate reads were filtered using Picard MarkDuplicates (2.18.11-SNAPSHOT) and further reads were also filtered based on MAPQ cut-off < 10.","Data Transformation - Normalized BigWig files for each sample were generated using makeUCSCfile program of HOMER Software. Total tags were normalized to default 1e7."],"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["NextSeq 550"],"study_type":["ChIP-seq"],"species":["Mus musculus"],"pubmed_authors":["Sunil Kumar Raghav"],"additional_accession":[]},"is_claimable":false,"name":"ChIP-seq of H3K27ac in Control and NCoR1 KD CD8α+ DCs at 0hr, 2hr and 6hr of TLR stimulation","description":"H3K27ac ChIP-seq was performed to identify active enhancers in different TLR activated control CD8α+ DCs as well as to understand the role of NCoR1 in regulating TLR specific enhacner activity using H3K27ac ChIP-seq performed in NCoR1 KD DCs. MutuDC1940 cell line were treated with TLR3 ligand poly I:C, TLR9 ligand CpG-B and combined stimulation of TLR9 , TLR3, and IFNg for 2hr and 6hr. Recombinant Anti-Histone H3 (acetyl K27) antibody was used to pull down chromatin regions having active enhacner mark in each unstimulated and stimulated condition.","dates":{"release":"2025-12-03T00:00:00Z","modification":"2026-05-27T06:00:49.011Z","creation":"2025-11-28T14:22:36.772Z"},"accession":"E-MTAB-10992","cross_references":{"ENA":["ERP185799"],"EFO":["EFO_0002944","EFO_0004170","EFO_0002692","EFO_0004917","EFO_0005518","EFO_0003816","EFO_0004184"]}}