|
| Status |
Public on Jan 15, 2019 |
| Title |
Tolerized KB5 (DST+antiCD154), rep1 |
| Sample type |
SRA |
| |
|
| Source name |
Tolerized KB5 CD8 T cells
|
| Organism |
Mus musculus |
| Characteristics |
strain/background: CBA/J
cell type: KB5 CD8 T cells
treatment: H2b antigens from C57BL/6 mice + anti-CD154
|
| Treatment protocol |
For donor-specific transfusion (DST), mice were injected IV with 10 million C57BL/6 splenocytes. For anti-CD154, mice were injected IP with 0.5 mg of the antibody. For LPS, mice were injected IP with 100 ug. Spleens were recovered from treated mice at 12 hours post treatment and stained with antibodies against the KB5 TCR and CD8. Cells were sorted to a 95% purity level.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was recovered from the purified cells. Extraction of total RNA from cultured mouse embryonic fibroblasts was performed using the RNeasy Mini Kit (Qiagen, Valencia, CA) according to the manufacturer's instructions. DNA was eliminated by on-column RNase-free DNase treatment. RNA quality was verified (RNA integrity number >8.5) using a 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
Messenger RNA was isolated, converted to cDNA, and amplified using the mRNA Purification Kit (Illumina, San Diego, CA) according to the manufacturer's instructions. An Illumina library was built and used to generate clusters on the Illumina flow cell according to the Illumina protocol and sequenced using a HiSeq 2000 (Illumina). Three independent libraries were examined for each condition.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
DSTMR1.CD8Tcell.rep1
Processed data file: mouse.gene.CD8Tcell.master_update_GEO_MATRIX.txt
|
| Data processing |
Three independent libraries were examined for each condition. The cDNA libraries were sequenced using the Illumina Hi-Seq 2000 platform with a paired-end 50-bp format. Reads from each sample were aligned to the mouse genome (UCSC genome browser mm10 build) using TopHat2 (Kim et al., 2013). The expression value for each gene was summarized using BedTools to count the number of aligned reads overlapping with exons of any annotated splice isoform in the RefSeq database (Quinlan et al., 2010). Gene expression values were quantified as reads per kilobase of gene length per million mapped reads (RPKM). Differentially expressed genes were further calculated using the DESeq R package (Anders and Huber, 2010) with the read count of the longest isoform of each gene as the input.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: mouse.gene.CD8Tcell.master_update_GEO_MATRIX.txt: Tab-delimited text file includes counts of raw reads that mapped to reads.
|
| |
|
| Submission date |
Oct 21, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Dr Yvonne Edwards, PhD |
| Organization name |
University of Massachusetts Medical School
|
| Department |
Program in Molecular Medicine
|
| Lab |
Bioinformatics Core
|
| Street address |
373 Plantation Street
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL13112 |
| Series (1) |
| GSE89030 |
Analysis of gene expression profiles from alloreactive TCR-Tg CD8 T cells during activation and induction of tolerance |
|
| Relations |
| BioSample |
SAMN05933396 |
| SRA |
SRX2256155 |
