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| Status |
Public on Apr 30, 2019 |
| Title |
MEL_PU.1_D0R2 |
| Sample type |
SRA |
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| Source name |
MEL
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| Organism |
Mus musculus |
| Characteristics |
cell type: MEL
chip antibody: PU.1
time: D0
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| Growth protocol |
Mouse erythroleukemia cells (MEL, ATCC, Manassas, VA) were grown in RPMI-1640 medium (Life Technologies, Carlsbad, CA) supplemented with penicillin/streptomycin (Life Technologies) and 10% FBS (VWR, Radnor, PA). MELs grown to a density of 1 million/ml were differentiated using 2% DMSO (VWR) added to the medium and the cells were grown for another 5 days to induce the expression of hemoglobin genes. At least 80-90% of cells expressed hemoglobin based on their color and benzidine/hydrogen peroxide staining result. Trypan Blue (VWR) staining was performed before harvesting the cells for each assay in order to make sure that at least 90% of the cells were viable. All experiments were done with 2 biological replicates.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Samples were extracted accoding to the Myers Lab ChIP-seq protocol v042211 (http://myers.hudsonalpha.org/documents/Myers%20Lab%20ChIP-seq%20Protocol%20v042211.pdf)
MEL PU.1 ChIP-seq libraries was made based on Myers Lab protocol v042211 (http://myers.hudsonalpha.org/documents/Myers%20Lab%20ChIP-seq%20Protocol%20v042211.pdf) using a rabbit polyclonal IgG PU.1 antibody (Santa Cruz Technology, sc-352, Lot#B0513, 200μg/ml). The quality of ChIP was validated by qPCR through the use of primers flanking known PU.1 binding sites (ex. SPI1, FCGR28, NRIH3) and known negative control sites (ex. MYOD1, ACTA1). The best two undifferentiated and differentiated MEL PU.1 ChIP replicates were constructed into Illumina sequencing libraries.
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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 |
PU.1 ChIP-seq reads were aligned to the mouse genome (mm9) with bowtie version 1.0.0 (Langmead et al. 2009) using the following settings: -S, –k 10, -m 10, --best, and --strata.
Peaks were called from the alignments using MACS2 version 2.0.10 (options: -p1e-3, -f BAM), and reproducible peaks were determined between the replicates using the Irreproducible Discovery Rate (IDR)(Li et al. 2011)
Reproducible replicate peaks that had IDRs < 0.05 and that were not present within mouse genomic blacklisted regions were used for downstream analyses.
Genome_build: mm9
Supplementary_files_format_and_content: Provided are 1) the unfiltered MACS2 peaks for each MEL PU.1 ChIP-seq replicate at day 0 (undifferentiated) and day 5 (differentiated with DMSO) in bed format (macs2_MEL_PU.1_d0r1_peaks.encodePeak, macs2_MEL_PU.1_d0r2_peaks.encodePeak, macs2_MEL_PU.1_d5r1_peaks.encodePeak, macs2_MEL_PU.1_d5r2_peaks.encodePeak), and 2) the IDR reproducible peaks between replicate sets that are filtered of blacklisted regions (MEL.PU1.peaks.d0.idrmerged.NOBLKlist.bed, MEL.PU1.peaks.d5.idrmerged.NOBLKlist.bed)
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| Submission date |
Apr 16, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Marissa Macchietto |
| E-mail(s) |
mmacchie@umn.edu
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| Organization name |
University of Minnesota, Minneapolis
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| Department |
Minnesota Supercomputing Institute
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| Street address |
117 Pleasant Street SE
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| City |
Minneapolis |
| ZIP/Postal code |
55455 |
| Country |
USA |
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| Platform ID |
GPL13112 |
| Series (1) |
| GSE67955 |
The landscape of global-long range interactions in mouse and human blood cell lines mediated by Yy1, GATA1, and CTCF during differentiation |
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| Relations |
| BioSample |
SAMN03487303 |
| SRA |
SRX997429 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1659604_macs2_MEL_PU.1_d0r2_peaks.encodePeak.txt.gz |
873.1 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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