|
| Status |
Public on Apr 30, 2024 |
| Title |
KO_Th2_R1 |
| Sample type |
SRA |
| |
|
| Source name |
Spleen and lymph nodes
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Spleen and lymph nodes
cell line: Primary LCMV TCRtg CD4+ Thy1.1+ cells
cell type: CD4 T cells
genotype: Tbx21-/-
treatment: Th2 conditions
|
| Treatment protocol |
Cells were cultured for two rounds under neutral (5 ng/ml IL-2 (Peprotech), 10 µg/ml anti–IL-4 (11B11), 10 µg/ml anti–IL-12 (C17.8), and 10 µg/ml anti–IFN-g (AN18.17.24)) or Th2 conditions (30 ng/ml IL-4 (Sigma-Aldrich), 5 ng/ml IL-2 (Peprotech), 10 µg/ml anti–IL-12 (C17.8), and 10 µg/ml anti–IFN-g (AN18.17.24)).
|
| Growth protocol |
Adoptively transferred WT, Tbx21+/-, Tbx21-/- LCMV‑TCRtg CD4+ Thy1.1+ cells from LCMV-challenged mice were isolated and MACS-sorted on day 10 of infection. Cells were cultured for two rounds under neutral (5 ng/ml IL-2 (Peprotech), 10 µg/ml anti–IL-4 (11B11), 10 µg/ml anti–IL-12 (C17.8), and 10 µg/ml anti–IFN-g (AN18.17.24)) or Th2 conditions (30 ng/ml IL-4 (Sigma-Aldrich), 5 ng/ml IL-2 (Peprotech), 10 µg/ml anti–IL-12 (C17.8), and 10 µg/ml anti–IFN-g (AN18.17.24)). On day 4 of the second round of stimulations the cells were harvested and processed for Chip-Seq.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from 106 cells using RNeasy mini kit (Qiagen) with a separate step for DNAse I digestion (Qiagen).
ERCC Spike-in control mix-1 (Life technologies) was added to each sample prior to processing (1μL of a 1:10 dilution of mix-1). The rRNA depletion and library construction was processed with the ScriptSeq Complete Gold Kit (Epicentre) according to the manufacturer’s instructions. The index for individual samples was added in order to multiplex 9 samples in each sequencing lane. The quality of libraries was assessed using a DNA1000 chip on the Bioanalyzer (Agilent) and the concentration was measured using the Qubit DNA assay. Sequencing was performed on the Illumina platform with HiSeq 2000 paired-end 100 bp sequence type.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Data processing |
#STAR v2.4.0j
STAR -- genomeDir <Directory > -- runThreadN 8 -- readFilesIn <Input file rep1 lane1 >,< Input file rep1 lane2 > <Input file rep2 lane1 >,< Input file rep2 lane2 > -- outFileNamePrefix <Prefix > -- outSAMtype BAM SortedByCoordinate
# HTSeq v0.6.1 python -m HTSeq.scripts.count -f bam <BAM Input file > <GTF file > > <Count file output >
#DESeq2 (v1.20) in R (v3.3.2) for vsd.txt file creation
Assembly: mm10
Supplementary files format and content: vsd.txt: tab-delimited text file with normalized count matrix (genes vs. samples)
Supplementary files format and content: .count files: tab-delimited text files with raw counts of the respective sample
|
| |
|
| Submission date |
Apr 29, 2024 |
| Last update date |
Apr 30, 2024 |
| Contact name |
Ahmed Nabil Hegazy |
| Organization name |
Charité - University Medicine Berlin
|
| Department |
Department of Gastroenterology, Infectious Diseases and Rheumatology
|
| Lab |
Hegazy Lab
|
| Street address |
Hindenburgdamm 30
|
| City |
Berlin |
| ZIP/Postal code |
12203 |
| Country |
Germany |
| |
|
| Platform ID |
GPL13112 |
| Series (1) |
| GSE266174 |
Plasticity and lineage commitment of individual Th1 cells are determined by stable T-bet expression quantities |
|
| Relations |
| BioSample |
SAMN41127338 |
| SRA |
SRX24402607 |
