|
| Status |
Public on May 20, 2022 |
| Title |
Lymph_node_endogenous_VDJ |
| Sample type |
SRA |
| |
|
| Source name |
Sorted CD4+ and CD4+CD8+ T cells from B16 melanoma resections
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Tumor infiltrating T cells
strain: RAG1- BW TRP-1 TCR
treatment: untreated
|
| Treatment protocol |
Cyclophosphamide (CTX) monohydrate mixed in sterile PBS was administered intraperitoneally as a single dose at 250mg/kg once tumors were established (1-2 weeks later). 1 day following CTX injection, 1x10^6 purified TRP1 CD4+ T cells were intravenously injected into the tail vein in 100ul PBS.
|
| Growth protocol |
B16F10 melanoma cells (2x105 cells) were implanted subcutaneously in 0.2ml of matrigel. Tumors were resected 11 days follow adoptive cell transfer and TRP1 (CD45.1) CD4+ and CD4+CD8+ TILS were FACs sorted.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Sorted T cells were stained with Trypan blue and the Countess II Automated Cell Counter (ThermoFisher) was used to assess both cell number and viability. Following QC, the single cell suspension was loaded onto Chromium Next GEM Chip K (10X Genomics PN 1000286) and GEM generation, cDNA synthesis, abd cDNA amplification.
Library preparation of 8,100 cells proceeded using the Chromium Next GEM Single Cell 5’ Kit v2 (10X Genomics PN 1000263) according to the manufacturer’s protocol. cDNA amplification included 13 cycles and 32ng of the material was used to prepare a sequencing library with 14 cycles of PCR. The indexed library was pooled equimolar and sequenced on a NovaSeq 6000 in a PE28/91 run using the NovaSeq 6000 SP Reagent Kit (100 cycles) (Illumina). 228 million paired reads were generated for the sample.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
10x VDJ
|
| Data processing |
Filtered gene expression matrices were generated using 10X CellRanger (version 3.1.0).
Paired single-cell VDJ sequences and annotations were generated using 10X Cellranger VDJ (version 3.1.0) and high confidence sequences were mapped to gene expression barcodes.
Assembly: mm10 (GENCODE vM23/Ensembl 98)
|
| |
|
| Submission date |
May 17, 2022 |
| Last update date |
May 20, 2022 |
| Contact name |
Bic MSKCC |
| Organization name |
Memorial SLoan-Kettering Cancer Center
|
| Street address |
1275 York Ave.
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10021 |
| Country |
USA |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE203182 |
Tumor-induced double positive T cells display distinct lineage commitment mechanisms and functions (1) |
| GSE203190 |
Tumor-induced double positive T cells display distinct lineage commitment mechanisms and functions |
|
| Relations |
| BioSample |
SAMN28488832 |
| SRA |
SRX15324776 |
