|
Status |
Public on Jun 14, 2018 |
Title |
S237_dog_KO4_CD8_p1_SC |
Sample type |
SRA |
|
|
Source name |
Tumor B16F10 melanoma
|
Organism |
Mus musculus |
Characteristics |
strain: C57BL/6 tissue: Tumor genotype/variation: IL27raKO
|
Treatment protocol |
Tumor infiltrating CD8+ T cells were isolated from WT and IL27ra KO B16F10 melanoma bearing mice.
|
Extracted molecule |
total RNA |
Extraction protocol |
5 µl lysis buffer comprised of Buffer TCL (Qiagen) plus 1% 2-mercaptoethanol. Plates were then spun down for one minute at 3000rpm and immediately frozen at -80°C. Cells were thawed and RNA was isolated with 2.2x RNAClean SPRI beads (Beckman Coulter Genomics) without final elution. The beads were then air-dried and processed immediately for cDNA synthesis. Samples were then processed using the Smart-seq2 protocol. This was followed by making a 25µl reaction mix for each PCR and performing 21 cycles for cDNA amplification. Then 0.25 ng cDNA from each cell and ¼ of the standard Illumina NexteraXT reaction volume were used in both the tagmentation and final PCR amplification steps
|
|
|
Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 2500 |
|
|
Description |
R3_rsem.genes.counts.matrix.TMM_normalized.FPKM
|
Data processing |
paired reads were mapped to mouse annotation mm10 using Bowtie (allowing a maximum of one mismatch in seed alignment, and suppressing reads that had more than 10 valid alignments). TPMs were computed using RSEM, and log2(TPM+1) values were used for subsequent analyses. filtered out low quality cells and cell doublets, maintaining for subsequent analysis the cells that had (1) 1,000-4,000 detected genes (defined by at least one mapped read), (2) at least 200,000 reads mapped to the transcriptome, and (3) at least 50% of the reads mapped to the transcriptome, ending with a total of 707 CD4+ and 825 CD8+ WT TILs and 376 CD4+ and 394 CD8+ IL27ra KO TILs. Genome_build: mm10
|
|
|
Submission date |
Apr 25, 2018 |
Last update date |
Jun 14, 2018 |
Contact name |
Asaf Madi |
E-mail(s) |
[email protected]
|
Organization name |
BWH
|
Department |
Neurology
|
Lab |
Ana Carrizosa Anderson
|
Street address |
60 Fenwood Road, BTM 10016F
|
City |
Boston |
State/province |
MA |
ZIP/Postal code |
02115 |
Country |
USA |
|
|
Platform ID |
GPL17021 |
Series (2) |
GSE113689 |
Single cell RNAseq of Wild Type and IL27ra KO Tumor infiltrating lymphocytes isolated from B16F10 melanoma batch 3 |
GSE113968 |
The induction and transcriptional regulation of the co-inhibitory gene module in T cells by IL-27 |
|
Relations |
BioSample |
SAMN08990635 |
SRA |
SRX4002276 |