title: GSM8974537: KPC63_IO; Mus musculus; RNA-Seq
library_construction_protocol: KPC63 tumors were established by injecting 1×10⁶ cells (1:1 PBS) subcutaneously into mouse hind legs. Tumor draining lymph nodes (TDLNs) were collected from three mice per treatment group, gently homogenized, and filtered through 40 μm filters in serum-supplemented medium. Tumor tissue underwent enzymatic digestion (HBSS/RPMI, 3% FetalClone II, DNAse I, Y-27632, Collagenase IV) for 45 min at 37°C. After filtration, RBC lysis, and resuspension in FetalClone II/10% DMSO, cells were stored in liquid nitrogen. For analysis, cells were revived in RPMI/5% FetalClone II, counted, and adjusted to 1000 cells/μL in 0.04% BSA/PBS. Samples were processed using 10x Chromium single cell 3' kit (V3 chemistry) to capture 10,000 cells, following standard protocol. Libraries were sequenced via NovaSeq to 50,000 reads per cell. Cells were loaded according to standard protocol of the Chromium single cell 5′ kit, capturing 10,000 cells (V3 chemistry). All subsequent procedures, including library construction, were performed according to the standard manufacturer's protocol. Single-cell libraries were sequenced on NovaSeq (Illumina) to a depth of 50,000 reads per cell