|
| Status |
Public on Aug 26, 2016 |
| Title |
Gfi1 GMP ChIP-Seq |
| Sample type |
SRA |
| |
|
| Source name |
Bone Marrow
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Granulocyte Macrophage Progenitors
strain: C57BL/6
library prep: ChIP Seq
chip antibody: Gfi1 antibody http://www.ncbi.nlm.nih.gov/pubmed/24141388
|
| Growth protocol |
Cells were harvested from bone marrow. Femurs, tibias and iliac crest were harvested immediately after euthanasia and put in cold PBS+2%FBS. Bones were crushed with mortar and pestle, filtered and washed in cold PBS+2%FBS, then enriched using CD117 MicroBeads on a Automacs Pro separator (Miltenyi, San Diego, CA). CD117+ cells were stained with CD16/32-PerCp-ef710 (clone 93), CD3-biotin (clone 145-2C11), CD4-biotin (clone RM4-5), CD8-biotin (clone 53-6.7), CD11b-biotin (clone M1/70), CD19-biotin (clone 6D5), Gr1-biotin (clone RB6-8C5), Ter119-biotin (clone Ter-119) CD45R-biotin (clone RA3-6B2), CD117-APC (clone 2B8), Sca-1-Pe-Cy7 (clone D7), CD34-BV421 (clone RAM34), CD115-BV605 (clone TR15-12F1 2.2), CD135-PE (A2F10.1), CCR3-FITC (clone 83101), SiglecF (clone E50-2440) and Gr1-FITC (clone RB6-8C5). Cell sorting was performed on MoFloXDP (Beckman Coulter, Brea, CA) or BD FACSAria II with a 100μm nozzleand subjected to flow cytrometric sorting of Lin- CD117+ Sca1- CD16/32+ CD34+ cells (GMP).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were fixed with 1% formaldehyde for 15 min and quenched with 0.125 M glycine. Lysates were sonicated with a microtip sonicator to shear the DNA. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation. For each ChIP reaction, 30 ug of chromatin was precleared with protein A agarose beads (Invitrogen). ChIP reaction was set up using precleared chromatin and antibody and incubated overnight at 4 C. Protein A agarose beads were added and incubation at 4 C was continued for another 3 hr. Immune complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment. Crosslinks were reversed by incubation overnight at 65 C, and ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation. After a final PCR amplification step, the resulting DNA libraries were quantified and sequenced on HiSeq 2500.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Lin- CD117+ Sca1- CD16/32+ CD34+
|
| Data processing |
ChIP-seq reads were aligned to the reference mm9 mouse genome using Bowtie2.
ChIP-seq peaks were called using the software Homer. Gene assignment was performed using the software GREAT (version 3.0.0), with the association rule of basal+extension: 5000 bp upstream, 1000 bp downstream, 100000 bp max extension, curated regulatory domains excluded.
Genome_build: mm9
Supplementary_files_format_and_content: Peak calling using HOMER in bed format. All parameters and commands are included (text file)
Supplementary_files_format_and_content: Tag statistics per chromosome
|
| |
|
| Submission date |
Jun 24, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
H. Leighton Grimes |
| E-mail(s) |
Lee.Grimes@cchmc.org
|
| Phone |
513-636-6089
|
| Organization name |
Cincinnati Childrens Hospital Medical Center
|
| Department |
Immunobiology
|
| Lab |
Grimes
|
| Street address |
3333 Burnet Ave. MLC 7038
|
| City |
Cincinnati |
| State/province |
OH |
| ZIP/Postal code |
45229 |
| Country |
USA |
| |
|
| Platform ID |
GPL17021 |
| Series (2) |
| GSE70237 |
Using single-cell RNA-Seq for unbiased analysis of developmental hierarchies (ChIP-seq) |
| GSE70245 |
Using single-cell RNA-Seq for unbiased analysis of developmental hierarchies |
|
| Relations |
| BioSample |
SAMN03792425 |
| SRA |
SRX1072347 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1721242_Gfi1_Tg_chr1.tags.tsv.gz |
2.0 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr10.tags.tsv.gz |
1.5 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr11.tags.tsv.gz |
1.6 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr12.tags.tsv.gz |
1.2 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr13.tags.tsv.gz |
1.4 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr13_random.tags.tsv.gz |
847 b |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr14.tags.tsv.gz |
1.2 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr15.tags.tsv.gz |
1.2 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr16.tags.tsv.gz |
1.1 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr17.tags.tsv.gz |
1.1 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr17_random.tags.tsv.gz |
115 b |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr18.tags.tsv.gz |
980.6 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr19.tags.tsv.gz |
799.9 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr1_random.tags.tsv.gz |
6.0 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr2.tags.tsv.gz |
2.1 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr3.tags.tsv.gz |
1.6 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr4.tags.tsv.gz |
1.7 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr4_random.tags.tsv.gz |
2.8 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr5.tags.tsv.gz |
1.7 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr6.tags.tsv.gz |
1.6 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr7.tags.tsv.gz |
1.5 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr7_random.tags.tsv.gz |
89 b |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr8.tags.tsv.gz |
1.4 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr8_random.tags.tsv.gz |
1.8 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr9.tags.tsv.gz |
1.4 Mb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chr9_random.tags.tsv.gz |
5.9 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chrM.tags.tsv.gz |
8.5 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chrUn_random.tags.tsv.gz |
21.8 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chrX.tags.tsv.gz |
740.7 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chrX_random.tags.tsv.gz |
3.0 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chrY.tags.tsv.gz |
6.8 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_chrY_random.tags.tsv.gz |
23.9 Kb |
(ftp)(http) |
TSV |
| GSM1721242_Gfi1_Tg_tagAutocorrelation.txt.gz |
27.3 Kb |
(ftp)(http) |
TXT |
| GSM1721242_Gfi1_Tg_tagCountDistribution.txt.gz |
7.0 Kb |
(ftp)(http) |
TXT |
| GSM1721242_Gfi1_Tg_tagInfo.txt.gz |
570 b |
(ftp)(http) |
TXT |
| GSM1721242_Gfi1_Tg_tagLengthDistribution.txt.gz |
308 b |
(ftp)(http) |
TXT |
| GSM1721242_Gfi1_peaks_5_10.txt.gz |
307.6 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
