|
| Status |
Public on Jun 14, 2018 |
| Title |
S155_cat_KO4_CD4_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 |
SAMN08990597 |
| SRA |
SRX4002195 |
