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| Status |
Public on Feb 28, 2024 |
| Title |
T-cell Resting 0h untreated rep3 [PAS-seq] |
| Sample type |
SRA |
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| Source name |
Sleen and lymph nodes
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
cell type: T-cell
cell status: Resting
biological replicate: Replicate 3
treatment: untreated
treatment time: 0h
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| Treatment protocol |
cells were incubated in the presence of 5,6-Dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) at a final concentration of 65µM or Triptolide (Sigma-Aldrich, Cat: T3652) at a final concentration of 25µM for 15min, 1h or 3h. At each time point (0, 15min, 1h and 3h), cells were collected, counted and total RNA was extracted from the same amount of cells at each time point. o monitor mRNA stability in conditions where mRNA translation is impaired, cells were incubated in the presence of either DRB or Triptolide and Cycloheximide (final concentration of 100µg.ml-1) or Harringtonine (final concentration of 2µg.ml-1) for 1 or 3h and total RNA extracted.
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| Growth protocol |
Primary CD4+ T cells were obtained from 6 week old C57BL/6J female mice. Briefly, the spleen as well as the inguinal, axillary, brachial, cervical and mesenteric lymph nodes were collected, followed by ficoll separation to remove red blood cells from splenocytes. CD4+ T cells were then purified by negative selection using the CD4+ T Cell Isolation kit (Myltenyi Biotec, Cat:130-104-454) following the manufacturer's protocol. Isolated cells were grown in RPMI medium supplemented with 10% foetal calf serum (FCS) and 50µM β-mercaptoethanol. CD4+ T cell activation was performed using magnetic beads coupled with CD3/CD28 antibodies (Thermo Fisher Scientific, Cat: 11452D) following the manufacturer’s protocol
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
total RNA was extrated using Trizol reagent, following manufacturer‘s procedure
High-throughput sequencing libraries were prepared as described in Heyer E et al., 2015 NAR
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| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
For PASeq
PAS_Resting_CD4_T_cells_R2
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| Data processing |
Library strategy: PAS-seq
Trimming with FASTX-Toolkits
Filtering with bowtie2.2.4 with -t –fast options against mouse 18S, 28S, 45S, 5S, and 5.8S rRNA, tRNAs
Mapping with TopHat 2 (v2.0.13) with --bowtie1 (bowtie version 1.1.1.0) --library-type fr-secondstrand --b2-sensitive -i 30 -m 1 -g 10 --max-coverage-intron 1000000 options
Count with Htseq with htseq-count -f sam -r pos -s yes -a 10 -m intersection-nonempty options
PASeq analysis was done as described in Ashar-Patel et.al, 2017 Sci Rep
Genome_build: mm10
Supplementary_files_format_and_content: coma-delimited text of each peak for each sample with count and genomic location
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| Submission date |
Oct 09, 2020 |
| Last update date |
Feb 28, 2024 |
| Contact name |
Emmanuel Labaronne |
| E-mail(s) |
elabaronne@gmail.com
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| Organization name |
LBMC
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| Lab |
RNA Metabolism in Immunity and Infection (RMI2)
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| Street address |
46 allée d'Italie
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| City |
Lyon |
| ZIP/Postal code |
69007 |
| Country |
France |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE159299 |
Translation-dependent and independent mRNA decay occur through mutually exclusive pathways that are defined by ribosome density during T Cell activation [PAS-Seq] |
| GSE159301 |
Translation-dependent and independent mRNA decay occur through mutually exclusive pathways that are defined by ribosome density during T Cell activation |
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| Relations |
| BioSample |
SAMN16403497 |
| SRA |
SRX9269832 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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