|
| Status |
Public on Nov 10, 2017 |
| Title |
GRO-Seq analysis of Wt mouse BMDM cells differentiated in the presence of MCSF (1hIL4) |
| Sample type |
SRA |
| |
|
| Source name |
bone marrow
|
| Organism |
Mus musculus |
| Characteristics |
treatment: cells differentiated in the presence of MCSF (1hIL4)
age: 8-12 weeks
tissue: bone marrow-derived macrophages
|
| Treatment protocol |
Mouse bone marrow-derived macrophages were treated with IL-4 (20 ng/ml; 1, hour).
|
| Growth protocol |
Isolation and differentiation were completed as described earlier (Daniel et al., 2014). Isolated bone marrow-derived cells were differentiated for 6 days in the presence of L929 supernatant.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Nuclei isolation was performed using buffer containing 10mM Tris-HCl, pH 7.4; 2mM MgCl2; 3mM CaCl2; 0.5% NP40; 10% Glycerol; 1mM DTT supplemented with protease inhibitors (prior to use) and RNase inhibitor (SUPERase-In, prior to use). Nuclear Run-On (NRO) reactions were performed in the presence of Br-UTP in duplicates containing ~10x106 nuclei and pooled for nascent RNA enrichment, using BrdU antibody coated agarose beads. Beads were washed and NRO RNA was eluted and quantified by Qubit fluorometer.
Approximately 100ng NRO RNA was used for end-repair with T4 PNK (3’-end de-phosphorylation) and Cap-Clip Acid Pyrophosphatase (de-capping). Libraries were prepared with NEBNext Small RNA Library Prep set for Illumina and sequenced on HiSeq 2500.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
mm_BMDM_1hIL4_GRO
nascent RNA
|
| Data processing |
Library strategy: GRO-Seq
alignment with bwa v0.7.10 using default parameters
bam generation with samtools v0.1.19 using default parameters
tag directory generation with homer v4.2 makeTagDirectory using -checkGC parameter
transcript calling with homer v4.2 findPeaks using-style groseq -tssSize 200 -minBodySize 300 -maxBodySize 3000 -tssFold 5 -bodyFold 2 -endFold 10 -fragLength 35
annotation was created with homer v4.2 annotatePeaks.pl using default parameters
Genome_build: mm10
Supplementary_files_format_and_content: annotation file
|
| |
|
| Submission date |
Nov 08, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Gergely Nagy |
| Organization name |
University of Debrecen
|
| Department |
Department of Biochemistry and Molecular Biology
|
| Street address |
Egyetem ter 1.
|
| City |
Debrecen |
| ZIP/Postal code |
4032 |
| Country |
Hungary |
| |
|
| Platform ID |
GPL17021 |
| Series (2) |
| GSE106703 |
Characterization of transcriptional and epigenetic changes during mouse alternative macrophage polarization [GRO-Seq] |
| GSE106706 |
Characterization of transcriptional and epigenetic changes during mouse alternative macrophage polarization |
|
| Relations |
| BioSample |
SAMN04868774 |
| SRA |
SRX3354458 |
