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Sample GSM2357674 Query DataSets for GSM2357674
Status Public on Jan 15, 2019
Title Tolerized KB5 (DST+antiCD154), rep3
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.rep3
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 SAMN05933393
SRA SRX2256157

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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