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| Status |
Public on Sep 02, 2021 |
| Title |
DL3A193_INPUT |
| Sample type |
SRA |
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| Source name |
CD19+ B cells
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| Organism |
Homo sapiens |
| Characteristics |
tissue: lymph node
disease state: diffuse large B cell lymphoma
antibody: input
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were formaldehyde cross-linked (1% final concentration) for 10 minutes in RPMI with 10% FBS. Quenching was performed by addition of 1M glycine (125 mM final concentration) with gentle mixing for 10 minutes at room temperature. Cells were washed with cold PBS and lysed in an SDS buffer (1% SDS, 10 mM EDTA, 50 mM Tris pH 8.0) with protease inhibitors before chromatin was fragmented using a Diagenode Bioruptor to an average size of less than 300 bp as determined using an Agilent Bioanalyzer 2100. Lysates were immunoprecipitated with the listed antibodies conjugated to Protein-A Dynabeads (5 ug antibody incubated with beads for 2-4 hours at 4°C) overnight at 4°C. The beads were then washed with the following buffers each for 3 minutes at 4°C - low salt buffer (150 mM NaCl, 50 mM Tris pH 8.0, 0.1% SDS), high salt buffer (500 mM NaCl, 50 mM Tris pH 8.0, 0.1% SDS), LiCl buffer (250 mM LiCl, 50 mM Tris pH 8.0, 0.5% Na deoxycholate, 1% NP40), and TE buffer (10 mM Tris pH 8.0, 1 mM EDTA). After washing, crosslinks were reversed overnight at 65°C in 250 uL elution buffer (1% SDS, 100 mM NaHCO3) followed by addition of proteinaseK and incubation at 56°C for 1 hour. DNA was purified with QIAquick PCR purification columns.
At least 3 ng of DNA was used to for indexed library preparation using Illumina TruSeq adapters and standard Illumina protocols. Fragments were size selected (average fragment-size: 200-400 bp for ChIP; 50-200 for FAIRE) using Agencourt AMPure XP beads prior to amplification. Samples were pooled (6-10 per lane) and sequenced.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Reads were aligned to hg19 with bowtie2 (v2.2.5) using default settings.
Reads in ENCODE blacklisted regions were removed with samtools (v1.9).
Peaks were called with MACS2 (v2.1.0.20150420) with the parameters --nomodel --shiftsize=150 (ChIP) or --nomodel --shiftsize=50 (FAIRE) and input controls.
DiffBind (v2.14.0) was used to derive consensus peak sets for each histone mark and accessible chromatin, requiring peaks to overlap in at least three samples for each ChIP-seq or FAIRE-seq assay in order to be merged and retained. The consensus peaksets for each assay were then concatenated and merged to derive a list of putative regulatory elements with bedtools2 (v2.29.2).
DiffBind was then used to determine differentially bound/accessible regions between sample groups for each assay. ChIPseeker (v1.22.0) was used to annotate peaks with hg19 UCSC knownGene annotations.
Genome_build: hg19
Supplementary_files_format_and_content: narrowPeak format files containing peaks for each sample as called by MACS2.
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| Submission date |
Feb 24, 2020 |
| Last update date |
Sep 02, 2021 |
| Contact name |
Jared Andrews |
| E-mail(s) |
jared.andrews@stjude.org
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| Organization name |
St. Jude Children's Research Hospital
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| Department |
Developmental Neurobiology
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| Street address |
262 Danny Thomas Pl
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| City |
Memphis |
| State/province |
Tennessee |
| ZIP/Postal code |
38105 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (2) |
| GSE145841 |
Key Super Enhancers Drive Tumor-Suppressing Transcription Feedback Programs in Mature B Cell Cancers (ChIP-seq, FAIRE-seq) |
| GSE145848 |
Key Super Enhancers Drive Tumor-Suppressing Transcription Feedback Programs in Mature B Cell Cancers |
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| Relations |
| BioSample |
SAMN14175969 |
| SRA |
SRX7795974 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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