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Status |
Public on Mar 18, 2019 |
Title |
Sample_Th1_4hrs_0612 |
Sample type |
SRA |
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Source name |
Murine Adult Spleen CD4 T Cells
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Organism |
Mus musculus |
Characteristics |
strain: C57BL/6 developmental stage: Murine Adult CD4 T cells cultured in vitro culture condition: Stimulated under Th1 for 4 hours genotype: Wildtype
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Treatment protocol |
CD4 Th cells were purified and set up in culture under three different Th differentiation conditions: Th0 condition was set up by treating the purified CD4 cells with anti-cd3e and anti-cd28; Th1 condition was set up with anti-cd3e, anti-cd28, anti-IL-4 and recombinant IL-12; and Th2 condition was set up with anti-cd3e, anti-cd28, anti-IL-12, anti-IFNG and recombinant IL-4.
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Extracted molecule |
total RNA |
Extraction protocol |
Murine CD4 positive cells were isolated from spleen cells by magnetic bead separation using EasySep mouse CD4+ cell negative selection kit (StemCell Technologies) according to the manufacturer’s instructions. The purity of isolated CD4+ cells was routinely exceeded 95%. For activation, CD4+cells were cultured at 5x106 cells/ml coated with 5µg/ml anti-CD3, and added 1µg/ml soluble anti-CD28 in complete RPMI and analysed at 4 and 20h. Th0, Th1 and Th2 skewing cultures, CD4+ cells were cultured on 5ug/ml plate bound anti-CD3 with were culture with 1µg/ml soluble anti-CD28 in complete RPMI. Th1 conditions the solution containing 5µg/ml anti-IL-4 and 10ng/ml rmIL-12. For Th2 cultures, 5µg/ml anti-IFNɣ; 20ng/ml rmIL-4, and 5µg/ml anti-IL-12 were added and, Cells were cultured at 37 °C under 5% CO2. Cells were harvested 72h and T-bet and Gata-3 were analysed through intracellular staining assay as described (Furmanski et al. 2013) (antibodies/proteins: eBioscience). RNA libraries were prepared for sequencing using standard Illumina protocols
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 2500 |
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Description |
Th1-4hours
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Data processing |
Base Calling was done using the RTA package from Illumina Alignment was done on the transcriptome using the Burrows-Wheeler Aligner (BWA) package Expression levels in Reads Per Million Kilobases (RPKM). Genome_build: GRCm38 Supplementary_files_format_and_content: comma separated file including RPKM values for each Sample.
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Submission date |
Jan 22, 2017 |
Last update date |
May 15, 2019 |
Contact name |
Tessa Crompton |
Organization name |
UCL
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Department |
Immunobiology
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Street address |
30 Guilford Street
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City |
London |
ZIP/Postal code |
WC1N 1EH |
Country |
United Kingdom |
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Platform ID |
GPL17021 |
Series (1) |
GSE93915 |
Genome Wide Transcriptional Modelling of a 24hour timecourse of T-helper cell differentiation |
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Relations |
BioSample |
SAMN06249346 |
SRA |
SRX2510467 |
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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