|
| Status |
Public on Jun 30, 2020 |
| Title |
Spleen_Endothelium_IFNg_RNA_1 |
| Sample type |
SRA |
| |
|
| Source name |
Murine structural cells
|
| Organism |
Mus musculus |
| Characteristics |
organ: Spleen
cell type: GP38negCD31pos
condition: IFNg
age: 8 to 12 weeks
strain: C57BL/6J
|
| Treatment protocol |
For the cytokine treatment experiments, mice were intravenously injected with 100 μg/kg of the following recombinant cytokines: IFN alpha, IFN beta, IL-3, IL-6, TGF-beta or TNF (all from Biolegend) and sacrificed 2 hours after cytokine injection.
|
| Growth protocol |
C57BL/6J mice, bred and maintained under specific pathogen free conditions at the Insti-tute of Molecular Biotechnology (IMBA) of the Austrian Academy of Sciences in Vienna, Austria. 8 to 12 week old mice (males only) were used for the steady state characterization
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Organs were digested with Accumax and structural cells isolated using FACS
For each individual assay, a maximum of 200 cells were sort-purified and deposited in 96-well plates con-taining 4 μl lysis buffer (1:20 solution of RNase Inhibitor (Clontech) in 0.2% v/v Triton X-100 (Sigma-Aldrich)), spun down, and immediately frozen at -80ºC. Reverse transcription and PCR were performed as described (Picelli et al., 2014). Library preparation was conducted on 1 ng of cDNA using the Nextera XT DNA Sample Preparation Kit (Illumina) followed by SPRI (Beckman Coulter) size selection. Sequencing was performed by the Biomedical Sequencing Facility at CeMM using the Illumina HiSeq 3000/4000 platform and the 50-bp single-end configuration.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Description |
SMART-seq sample of Spleen GP38negCD31pos cells
|
| Data processing |
Adapter trimming with trimmomatic (version 0.32) with settings: SLIDINGWINDOW:4:1 MAXINFO:16:0.40 MINLEN:30
Genome alignment using HISAT (2.1.0) with default parameters
Filtered reads to primary alignments (excluding secondary alignments) to exons (defined by Ensembl, genome-build GRCm38.p6 in the GENCODE Basic set) using the GenomicAlignments package in R (3.2.3)
Counted reads in genes over all samples using the package GenomicAlignments in R (3.2.3)
Normalized to counts per million (CPM) and transformed to log2(CPM) using Limma Voom in R
Regressed out immune cell signatures using Limma removeBatchEffect in R
Genome_build: mm10
Supplementary_files_format_and_content: Matrix of read counts (CSV), read counts of each gene for each sample
Supplementary_files_format_and_content: Matrix of log2(CPM) values (CSV), log2(CPM) after regressing out immune signatures of each gene
|
| |
|
| Submission date |
Mar 06, 2020 |
| Last update date |
Jun 30, 2020 |
| Contact name |
Christoph Bock |
| E-mail(s) |
cbock@cemm.oeaw.ac.at
|
| Organization name |
CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences
|
| Street address |
Lazarettgasse 14
|
| City |
Vienna |
| ZIP/Postal code |
1090 |
| Country |
Austria |
| |
|
| Platform ID |
GPL21103 |
| Series (2) |
| GSE134663 |
Structural cells are key regulators of organ-specific immune response |
| GSE145820 |
Structural cells are key regulators of organ-specific immune response |
|
| Relations |
| BioSample |
SAMN14322834 |
| SRA |
SRX7866719 |
