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| Status |
Public on Sep 19, 2020 |
| Title |
Vec6-BMP-plus-PU1-ChIP |
| Sample type |
SRA |
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| Source name |
K562
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| Organism |
Homo sapiens |
| Characteristics |
cell type: immortalized erythroleukemia cells (empty control)
purification target: PU1
antibody manufacturer: Santa Cruz
antibody catalog id: sc352X
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| Treatment protocol |
hrBMP4 25ng/ml 2hrs
25ng/ml hrBMP4 for 2hrs prior to collection
Whole cell extracts were sonicated to solubilize the chromatin and target bound fragments were isolated with antibody
Human CD34+ cells, isolated from the peripheral blood of granulocyte colony-stimulating factor mobilized healthy volunteers, were used.Briefly the cells were expanded in StemSpan medium (Stem Cell Technologies Inc.) supplemented with StemSpan CC100 cytokine mix (Stem Cell Technologies Inc.). For studying differentiated cells after day 6 of expansion, cells were reseeded in differentiation medium (StemSpan SFEM Medium with 2% P/S, 20 ng/ml SCF, 1 U/ml Epo, 5 ng/ml IL-3, 2 mM dexamethasone, and 1 mM b-estradiol). K562 cells were grown and maintained in RPMI base media with FBS, Pen/Strep. G418 was used to select positive clones.A72:A74
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| Growth protocol |
Human CD34+ cells, isolated from the peripheral blood of granulocyte colony-stimulating factor mobilized healthy volunteers, were used.Briefly the cells were expanded in StemSpan medium (Stem Cell Technologies Inc.) supplemented with StemSpan CC100 cytokine mix (Stem Cell Technologies Inc.). For studying differentiated cells after day 6 of expansion, cells were reseeded in differentiation medium (StemSpan SFEM Medium with 2% P/S, 20 ng/ml SCF, 1 U/ml Epo, 5 ng/ml IL-3, 2 mM dexamethasone, and 1 mM b-estradiol). K562 cells (over expressing PU1) were grown and maintained in RPMI base media with FBS, Pen/Strep. G418 was used to select positive clones. K562 cells (with PU1 knockdown) were grown and maintained in RPMI base media with FBS, Pen/Strep. puromycin was used to select positive clones.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
The chromatin extracts containing DNA fragments with an average size of 500 bp were immunoprecipitated using different antibodies. Purified immunoprecipitated DNA were prepared for sequencing according to a modified version of the Solexa Genomic DNA protocol. Fragmented DNA was end repaired and subjected to 18 cycles of LM-PCR using oligos provided by Illumina. Amplified fragments between 150 and 300bp (representing shear fragments between 50 and 200nt in length and ~100bp of primer sequence) were isolated by agarose gel electrophoresis and purified.
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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 |
Aligned using bowtie with configuration -p 4 --best -k 1 -m 1 --sam -l 40
Genome_build: hg19
Supplementary_files_format_and_content: WIG files(s) represent counts of aligned reads within 50 bp bins with each read being extended 200 in the direction of alignment. Counts are in reads-per-million and floored at 0.1
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| Submission date |
Sep 18, 2020 |
| Last update date |
Sep 20, 2020 |
| Contact name |
Leonard Zon |
| E-mail(s) |
zon@enders.tch.harvard.edu
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| Organization name |
Boston Children's Hospital
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| Department |
Oncology/Hematology
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| Street address |
1 Blackfan Circle
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE104574 |
Transcriptional Signaling Centers Regulate Erythroid Gene Expression and are Disrupted in Common Variations of Human Red Blood Cell Traits |
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| Relations |
| BioSample |
SAMN16212682 |
| SRA |
SRX9151994 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4795524_AC3_hg19_peaks.narrowPeak.gz |
470.4 Kb |
(ftp)(http) |
NARROWPEAK |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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