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| Status |
Public on Aug 07, 2024 |
| Title |
polyclonal CD8 cells TCR seq |
| Sample type |
SRA |
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| Source name |
Tumor and lymph node
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| Organism |
Mus musculus |
| Characteristics |
tissue: Tumor and lymph node
cell line: B16
cell type: CD8
library type: TCR
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| Extracted molecule |
other |
| Extraction protocol |
Single-cell libraries were prepared from sorted live CD8+ T cells derived from either polyclonal mice or CD8+ pMEL cells from an antigen-specific system, utilizing Chromium Single Cell 5’ Reagent (v1 Chemistry) or Chromium Next GEM Single Cell 5’ Reagent (v2 Chemistry Dual Index) immediately post-cell isolation.
Sample multiplexing was achieved via CITEseq, wherein cells were stained with TotalSeq-C anti-mouse hashtag antibodies and additional surface flow cytometry antibodies (TCRβ, CD8). Sorted cells, tagged with uniquely barcoded TotalSeq-C antibodies, were pooled and loaded into parallel lanes for droplet generation, targeting a recovery of 2,000 cells per sample. After droplet partitioning, cells underwent lysis and reverse transcription within droplets.
cDNA was isolated, bulk amplified, and selected with SPRIselect beads. Adaptors were ligated, sample indices were added through PCR, and samples underwent another selection using SPRIselect beads. Library concentration was determined via PCR with KAPA DNA Quantification.
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| Library strategy |
OTHER |
| Library source |
other |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
TCR seq
TCR library, read 1 contains barcode and UMI information, and read 2 containsTCR sequences
polyclonal_TCR
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| Data processing |
After library preparation and quantification, samples were pooled and sequenced on a NovaSeq 6000 using S1 v1.5 kits (150 cycles), with sequencing parameters adjusted for v1 and v2 Chemistry.
Demultiplexing, barcode processing, gene counting, and aggregation were conducted using Cell Ranger software version 6.1.2 and CITE-seq-Count version 1.4.4.
Assembly: mm10
Supplementary files format and content: CellRanger output files: polyclonal_GEX_barcodes.tsv.gz, polyclonal_GEX_features.tsv.gz, polyclonal_GEX_matrix.mtx.gz
Supplementary files format and content: CellRanger output files: polyclonal_TCR_filtered_contig_annotations.csv
Supplementary files format and content: CellRanger output files: pMEL_GEX_barcodes.tsv.gz, pMEL_GEX_features.tsv.gz, pMEL_GEX_matrix.mtx.gz
Supplementary files format and content: CITE-seq-Count output files: polyclonal_HTO_barcodes.tsv.gz, polyclonal_HTO_features.tsv.gz, polyclonal_HTO_matrix.mtx.gz
Supplementary files format and content: CITE-seq-Count output files: pMEL_HTO_barcodes.tsv.gz, pMEL_HTO_features.tsv.gz, pMEL_HTO_matrix.mtx.gz
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| Submission date |
Mar 28, 2024 |
| Last update date |
Aug 07, 2024 |
| Contact name |
Jian Cui |
| E-mail(s) |
cuij@pitt.edu
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| Organization name |
University of Pittsburgh
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| Department |
Immunology
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| Lab |
Vignali Lab
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| Street address |
5051 Centre Avenue
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| City |
Pittsburgh |
| State/province |
PA |
| ZIP/Postal code |
15261 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE262740 |
LAG3 and PD1 synergize on CD8+ T cells to drive T cell exhaustion and hinder autocrine IFN gamma-dependent anti-tumor immunity |
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| Relations |
| BioSample |
SAMN40648465 |
| SRA |
SRX24095536 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8174731_polyclonal_TCR_filtered_contig_annotations.csv.gz |
1.5 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
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