GSM9449372: Brain, GL261-CAR, oAd, CP3; Mus musculus; RNA-Seq
Mice bearing intracranial GL261‑CAR tumors were treated intratumorally with PBS (control), oAd, or oAd‑mCD47/PD‑L1‑mIL‑12 (1 × 10^9 pfu). Tumors were harvested three days after the second viral injection and dissociated into single‑cell suspensions using the SeekMate Tissue Dissociation Reagent Kit A Pro (SeekGene, K01801‑30) at 37 °C for 15 min. Erythrocytes were lysed with an erythrocyte removal reagent (Solarbio, R1010). Cell count and viability were determined using a SeekMate Tinitan Fluorescence Cell Counter (SeekGene, M002C) with AO/PI staining. Debris and dead cells were removed by sequential filtration through a 40‑μm cell strainer and a dead‑cell removal kit (Miltenyi Biotec, #130‑090‑101). Cells were washed twice in RPMI‑1640 medium and resuspended at 1 × 106 cells/mL. Single‑cell RNA‑seq libraries were constructed using the SeekOne Digital Droplet Single‑Cell 3′ Library Preparation Kit (SeekGene, #K00202) according to the manufacturer's protocol. Libraries were sequenced on an Illumina NovaSeq X Plus platform with 150‑bp paired‑end reads. Raw sequencing reads were processed with fastp (v0.23.2) to trim adapters and remove low‑quality bases. Cellular barcodes were demultiplexed and reads were aligned to the mouse reference genome (mm10) using SeekSoul tools (SeekGene, v1.2.1).