 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jul 28, 2014 |
| Title |
Thymus_H3K4me2 |
| Sample type |
SRA |
| |
|
| Source name |
Thymocytes
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
age: 4 weeks
tissue: thymus
cell type: Thymocytes
genome/variation: wild-type
antibody: H3K4me2, antibody vendor, cat#, lot #: Millipore, 07-030, DAM1724042
|
| Treatment protocol |
To expand erythroid cells, granulocytes, and MKs, cultures were supplemented, respectively, with Stem Cell Factor (SCF, R&D Systems, 0.02 mg/10 mL) and erythropoietin (EPO, R&D Systems, 0.02 mg/10 mL); SCF and granulocyte-colony stimulating factor (G-CSF, R&D Systems, 0.05 mg/10 mL); or thrombopoietin (TPO, 1/100 dilution of producer cell supernatant [Villeval, 1997 #2788]). To enrich hematopoietic progenitors, short-term cultures were supplanted with SCF and interleukin-3 (R&D Systems, 0.05 mg/10 mL). Progenitors were harvested after 1 day of culture, IMM lineages after 2 days, and MAT cells after 4 (erythroid, granulocyte) or 5 days (MKs).
|
| Growth protocol |
Thymocytes were isolated from the thymus of 4 week old mice. Cells were immediately subjected to H3K4me2 ChIP-seq.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Individual lineages were isolated by depleting other lineages using TER119, GR1, CD41, and CD11b (B-D Pharmingen) monoclonal antibodies (mAb) and immunomagnetic Dynabeads (Invitrogen) or by positive selection using GR1 or TER119 mAb for flow cytometry. MK isolations included additional passage over triple (4%, 3%, 1.5%) gradients of bovine serum albumin, with collection of the supernatant (IMM cells) or pellet (MAT cells). Cell isolations were monitored by viewing May-GrĂĽnwald-Giemsa stains of cytocentrifuged preparations with an Olympus BX41 microscope and by flow cytometry using the above-listed mAb. Chromatin marks were pulled down with H3K4me2 or H3K27ac antibodies after MNase digestion or crosslinking + sonication of the cells. Transcription factors were pulled down with the respective antibodies after crosslinking + sonication of the cells
At least 20 ng of precipitated or input DNA from pools of at least 3 replicates was processed for deep sequencing using different library preparation kits for IMM or MAT cells (TruSeq, Illumina) and progenitors (TruPLEX-FD, Rubicon Genetics), following the manufacturers’ instructions. ChIP-seq libraries were sequenced using Illumina HiSeq2000 and tags were mapped to mouse genome build mm9.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
ChIP-seq reads were aligned to the mm9 genome assembly using bowtie with -m 1 and default parameters
Uniquely aligned reads were filtered using SICER v1.1 to keep up to 2 copies at each genomic location
Wiggle files for histone modifications were generated using NPS with default parameters
Wiggle files for progenitor cells' H3K4me2 and transcription factors were generated using MACS v1.4
Genome_build: mm9
|
| |
|
| Submission date |
Nov 07, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Chongzhi Zang |
| Organization name |
University of Virginia
|
| Street address |
P. O. Box 800717
|
| City |
Charlottesville |
| State/province |
VA |
| ZIP/Postal code |
22908 |
| Country |
USA |
| |
|
| Platform ID |
GPL13112 |
| Series (1) |
| GSE42110 |
Epigenetic and transcriptional control in hematopoietic development and lineage differentiation |
|
| Relations |
| SRA |
SRX203218 |
| BioSample |
SAMN01803704 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1032648_Thymus_H3K4me2.wig.gz |
39.0 Mb |
(ftp)(http) |
WIG |
| GSM1032648_Thymus_H3K4me2_2copy.bed.gz |
235.8 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
