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| Status |
Public on Feb 08, 2018 |
| Title |
RNA_VitDex_2hr_BR2_TR1 |
| Sample type |
SRA |
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| Source name |
VitDex,6hr,in vitro
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| Organism |
Mus musculus |
| Characteristics |
genotype: wildtype
background strain: C57BL/6
tissue: spleen
cell type: CD4+ T cell
cd4+ t cell subset: VitDex
time point: 6hr
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| Treatment protocol |
Cultured with Vitamin D3, Dexamethasone, anti-IFNg, anti-IL-12p40, anti-IFNg and anti-IL-4 for 7d, re-stimulated with anti-CD3/CD28 for 6h.
CD4+ T cells were differentiated as described where biological replicates [BR] refer to different experiment, technical replicates [TR] are different culture wells within the same biological replicate. Some samples were sequenced in two runs. These samples are identified in the 'description' field and include two SRR per sample.
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| Growth protocol |
CD4+ T cells were negatively enriched from spleen using goat anti-MHCII, anti-CD8 and anti-B220 and anti-goat beads (Quiagen), followed by staining in PBS with 5% FCS for 20 min at 4C with anti- anti-CD8-FITC, anti-CD4-eFluor450, anti-CD25-APC, anti-CD62L-PECy7, anti-CD44-PE (eBioscience). Live (propidium iodide negative) CD8-CD4+CD25-CD62L+CD44lo T cells were sorted on a MoFloTM XDP cytometer (Beckman Coulter). Sorted cells were activated with anti-CD3 (5 ug/ml) and anti-CD28 (2 ug/ml) and cultured in RPMI in the presence of Vitamin D3 (4x10-8M, Enzo Life Sciences), Dexamethasone (1x10-8M, Sigma-Aldrich), anti-IFNg (Harlan), anti-IL-12 (gift from Dr. G. Trinchieri) and anti-IL-4 (gift from DNAX) at 10 ug/ml for 21 days with anti-CD3/28 (2 ug/ml) restimulation in the presence of the above on day 7 and day 14. On day 21, cells were harvested and restimulated with anti-CD3 (2 ug/ml) and anti-CD28 (2ug/ml) for 6h.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Extraction of total RNA was carried out using the RNeasy Mini Kit (Qiagen) according to manufacturer’s instructions.
Libraries were prepared with TruSeq RNA Sample Preparation Kit V2 (Illumina) according to the manufacturer’s instructions.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Alignment of reads to the mouse transcriptome (mm10) and absolute quantification of the genes was performed in Strand NGS (version 2.0), merging files from the different sequencing runs when available, using default parameters (95% identity, max 5% gaps, 1 read only if duplicated, ignoring reads with more than 5 matches) and guided by RefSeq annotations (2013.04.01).
Genome_build: mm10
Supplementary_files_format_and_content: bigwig
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| Submission date |
Nov 02, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Leona Gabrysova |
| E-mail(s) |
Leona.Gabrysova@crick.ac.uk
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| Phone |
+442037961441
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| Organization name |
The Crick Institute
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| Lab |
Immunoregulation and Infection Lab
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| Street address |
1 Midland Road
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| City |
London |
| ZIP/Postal code |
NW1 1AT |
| Country |
United Kingdom |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE106463 |
The transcription factor c-Maf is a positive regulator of IL-10 with context-specific roles in CD4+ T cell effector function and in vivo consequences [RNA-seq in vitro] |
| GSE106464 |
The transcription factor c-Maf is a positive regulator of IL-10 with context-specific roles in CD4+ T cell effector function and in vivo consequences |
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| Relations |
| BioSample |
SAMN07967579 |
| SRA |
SRX3353707 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2837981_VitDex_2hr_BR2_TR1-All_Aligned_Reads.bw |
4.6 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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