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| Status |
Public on May 13, 2019 |
| Title |
Old_33_H3 |
| Sample type |
SRA |
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| Source name |
Bone marrow
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| Organism |
Homo sapiens |
| Characteristics |
cell type: HSCe (CD34+, CD38-,Lineage-)
donor age: 73
donor sex: M
antibody: Abcam ab10799, lot #GR275925-1
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| Treatment protocol |
Miltenyi MACS magnetic bead purification was used to enrich frozen bone marrow MNC for CD34+ cells. The CD34+ fraction was then stained with CD2-TC, CD3-TC, CD4-TC, CD7-TC, CD8-TC, CD10-TC, CD11b-TC, CD14-TC, CD19-TC, CD20-TC, CD56-TC, and Glycophorin A-TC followed by CD34-APC, CD38-PE-Cy7, CD123-PE, CD45RA-FITC and DAPI. Using a BD FACS SORP Aria-Ilu HSCe (DAPI-,Lin-, CD34+, CD38-) were FACS sorted. HSCe were sorted into 1mL IMDM 20% FBS. 10,000-15,000 HSCe were used per immunoprecipitation.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Micro-ChIPseq was performed using the True MicroChIP (Diagenode, #C01010130) kit and 0.5 ug H3 (Abcam ab10799, lot #GR275925-1) per immunoprecipitation
ChIP-seq libraries were prepared using the V1 MicroPlex Library Preparation kit (Diagenode, #C05010011). For the PCR amplification, a total of 16 amplification cycles was used. Libraries were purified using a 1:1 Ampure bead cleanup and eluted in 16 uL of Tris-HCl ph 8.0. Libraries were multiplexed and sequenced using a NextSeq 500 with 75bp single end sequencing.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Description |
H3 pulldown
O3_H3_FE.bw
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| Data processing |
FastQC was run all samples. Samples with a sequencing duplication rate of >90% were discarded.
Adapters and reads of less than 25 basepairs were removed using cutadapt (version 1.12).
Reads were aligned to the hg19 genome using bowtie2 (version 2.0.5).
Reads were filtered to retain only those that were aligned and had one alignment (!XS:i:N) samtools (version 1.2)
Unique mapped reads for each IP type for each age group were pooled using samtools merge (version 1.2)
Deeptools bamCompare with --scaleFactorsMethod readCount and --ratio log2 was used to generate a bigwig file containing the log2 enrichment of H3 compared to Input
Genome_build: hg19
Supplementary_files_format_and_content: The processed file is a bigwig file containing the log2(H3/Input), normalized by read count.
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| Submission date |
Nov 02, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Maria E. Figueroa |
| E-mail(s) |
[email protected]
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| Phone |
305-243-7333
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| Organization name |
University of Miami Miller School Of Medicine
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| Department |
Human Genetics
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| Street address |
1501 NW 10th Ave, BRB 742F
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| City |
Miami |
| State/province |
Florida |
| ZIP/Postal code |
33136 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE104408 |
Aging Human Hematopoietic Stem Cells Manifest Profound Epigenetic Reprogramming of Enhancers That May Predispose to Leukemia |
| GSE106422 |
Aging Human Hematopoietic Stem Cells Manifest Profound Epigenetic Reprogramming of Enhancers That May Predispose to Leukemia (H3 ChIPseq) |
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| Relations |
| BioSample |
SAMN07972747 |
| SRA |
SRX3358471 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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