biostudies-arrayexpressShunsuke TakasugaMus musculushttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-15883Processed single-cell datasets from Pdcd1-CreERT2 × Rosa26-tdTomato fate-mapping mice bearing subcutaneous LLC allografts and treated with CTX, anti-PD-1, or both. We provide count matrices, cell-level annotations, and per-cell TCR clonotypes; raw reads for transcriptome, sample-tag, and TCR libraries are deposited under ENA BioProject PRJEB100721. This resource profiles PD-1 lineage cells, highlighting therapy-dependent changes in progenitor-exhausted CD8 T cells and TCR clonotype dynamics.biostudies-arrayexpressSample Treatment - Pdcd1-Cre-ERT2 mice were generated by CRISPR/Cas9-mediated insertion of a P2A–Cre-ERT2 cassette at the 3′ end of the endogenous Pdcd1 coding region. For fate mapping of PD-1+ cells, Pdcd1-Cre-ERT2 mice were crossed with Rosa26-loxP-STOP-loxP-tdTomato mice (JAX #007914) to generate Pdcd1fm animals. Female Pdcd1fm mice aged 8–11 weeks were used. A total of 6 × 10^4 Lewis lung carcinoma (LLC) cells in 100 µL PBS were injected subcutaneously into the left flank. Tamoxifen was administered to induce tdTomato expression. On day 12 post-implantation, mice were randomized into four groups with matched tumor volumes (control [NC], CTX, ICI, and CTX+ICI) and received the first treatment. CTX (100 mg/kg) and/or anti–PD-1 antibody (clone RMP1-14, 400 µg per mouse) were administered intraperitoneally on day 12 and again on day 19.Nucleic Acid Extraction - No separate bulk nucleic-acid extraction was performed. Cells were loaded onto the BD Rhapsody cartridge; lysis and capture on barcoded beads occurred in situ during reverse transcription. Downstream libraries (WTA, Sample-Tag, TCR) were prepared directly from bead-captured cDNA per the manufacturer’s instructions.Sample Collection - Tumors were harvested on day 23 post-implantation (11 days after the first treatment). Dorsal tumors were excised, finely minced, and incubated at 37 °C, 200 rpm for 45 min in 20 mL digestion buffer (RPMI + 2% FBS, 10 mg collagenase IV, 1 mg DNase). Cell suspensions were filtered through 70-µm strainers. RBCs were removed by brief hypotonic lysis, and leukocytes were enriched using a 40% Percoll procedure. Dead cells were excluded using Fixable Viability Dye eFluor 506.Library Construction - Single-cell whole-transcriptome (WTA) libraries were prepared using the BD Rhapsody workflow. Read structure: R1 = cell barcode(s) + UMI; R2 = cDNA insert; i7 = sample index. In parallel, Sample-Tag (HTO) libraries were generated (R1 = barcode + UMI; R2 = HTO insert) after labeling each condition with BioLegend TotalSeq-A hashtags A0301–A0304 and pooling. TCR α/β (TRA/TRB) amplicon libraries were prepared using the BD Rhapsody Mouse TCR Amplification Kit with PCR enrichment and purification. Libraries were quantified (KAPA qPCR) and sized (MultiNA/Bioanalyzer).Sequencing - A short iSeq run was used for pool-balance QC. Final pooled WTA, HTO, TCR libraries were sequenced on an Illumina NovaSeq 6000 (paired-end; i7 index). Exact cycle settings are provided in the ENA Run records. Raw FASTQs are deposited in ENA under BioProject PRJEB100721.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - FASTQs were processed with the BD Rhapsody pipeline to extract cell barcodes/UMIs and align to GRCm39 (Ensembl rel-107) with UMI collapsing to gene-by-cell matrices (Matrix Market). HTO counts were used for demultiplexing and multiplet removal. Count matrices were analyzed in Seurat v5 (R): SCTransform v2 normalization (defaults, no covariates), PCA, k-NN graph construction, graph-based clustering (Seurat defaults), and UMAP embedding. Cluster 15 (tumor cells) was excluded from downstream analyses. Processed outputs include matrix.mtx.gz, features.tsv.gz, barcodes.tsv.gz, and cell_metadata.tsv.UnknownTranscriptomicsGenomicsProteomicsIllumina NovaSeq 6000RNA-seq of coding RNA from single cellsMus musculusShunsuke TakasugafalseSingle-cell transcriptome and TCR repertoire of PD-1 fate-mapped immune cells during cyclophosphamide and PD-1 blockade in syngeneic subcutaneous LLC tumor allograftsProcessed single-cell datasets from Pdcd1-CreERT2 × Rosa26-tdTomato fate-mapping mice bearing subcutaneous LLC allografts and treated with CTX, anti-PD-1, or both. We provide count matrices, cell-level annotations, and per-cell TCR clonotypes; raw reads for transcriptome, sample-tag, and TCR libraries are deposited under ENA BioProject PRJEB100721. This resource profiles PD-1 lineage cells, highlighting therapy-dependent changes in progenitor-exhausted CD8 T cells and TCR clonotype dynamics.2026-03-11T00:00:00Z2026-03-11T06:52:49.263Z2025-10-27T21:34:02.971ZE-MTAB-15883ERP183151EFO_0002944EFO_0004170EFO_0005684EFO_0005518EFO_0003816EFO_0004184EFO_0003969