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| Status |
Public on Jan 22, 2017 |
| Title |
WT_PolII_rep_1 |
| Sample type |
SRA |
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| Source name |
Thymus
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| Organism |
Mus musculus |
| Characteristics |
strain: C57Bl/6
genotype: Aire+/+
cell type: MECs
antibody: PolII
antibody vendor/catalog: Covance/MMS-128P
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were isolated by tissue digestion and fluorescence activated cell sorting for viable CD45-Ly51loMHCIIhi as MECs.
Chromatin IPed with specific Abs was eluted from the Protein-G beads, treated with 1µg DNase-free RNase (Roche) for 30 min at 37°C and with Proteinase K (Roche) for 2 hrs at 37°C followed by reverse cross-linking by leaving the plate at 65°C overnight. DNA from reverse cross-linked material was purified with SPRI beads (Agencourt AMPure XP beads, Beckman Coulter); and sequential steps of end-repair, A-base addition, adaptor-ligation and PCR amplification (15 cycles) were performed to prepare the ChIP-seq library for each sample.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Short reads (50bp, single end) were aligned to the mouse reference genome (mm10) using bowtie aligner version 2.2.4
Reads with multiple alignments were removed with samtools (v1.1) and de-duplicated with picard (v1.130)
To identify peaks from ChIP-seq reads, we used the HOMER package makeTagDirectory followed by the findPeaks command with the ‘histone’ parameter. Peaks displaying 4-fold enrichment and poison P-value of 1x10-4 against background IgG ChIP were considered significant and were used for further analysis.
To visualize individual ChIP-seq data on Integrative Genomics Viewer (IGV), we converted bam output files from picard into normalized bigwig format using the bamCoverage function in deepTools (v1.6) with options – fragmentLength 200 –normalizeUsingRPKM
Genome_build: mm10
Supplementary_files_format_and_content: bigwig
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| Submission date |
Dec 19, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
CBDM Lab |
| E-mail(s) |
cbdm@hms.harvard.edu
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| Phone |
617-432-7747
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| Organization name |
Harvard Medical School
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| Department |
Microbiology and Immunobiology
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| Lab |
CBDM
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| Street address |
77 Avenue Louis Pasteur
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE92597 |
Aire, guardian of immunological tolerance, binds to and activates super-enhancers [ChIP-seq] |
| GSE92654 |
Aire, guardian of immunological tolerance, binds to and activates super-enhancers |
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| Relations |
| BioSample |
SAMN06167704 |
| SRA |
SRX2437687 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2433218_WT_PolII_rep_1.with.adapter.mapped_1align.sorted.uniq.RPKM.bigWig |
18.6 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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