title: GSM9373025: TAM_cGAMPnp2; Mus musculus; RNA-Seq
library_construction_protocol: Tumour tissues were cut into small pieces and digested at 37°C for 30 min by incubation with collagenase type 1A (1.5 mg/mL) and hyaluronidase (1.5 mg/mL) in DMEM. The cells were then filtered with 40 um cell strainers followed by a centrifugation at 3000 rpm for 10 min. Following washes with 1% BSA in PBS, red blood cells were removed using 1× ACK lysis buffer. Dead cells were subsequently deleted using a dead cell removal kit and MS columns, and the resulting viable single-cell suspensions were used for downstream single-cell RNA sequencing. Single cells were processed through the GemCode Single Cell Platform using the Chromium Fixed RNA Kit, Mouse Transcriptome (10X Genomics, Pleasanton) according to  the manufacturer's protocol. In brief, Cell suspensions were fixed with paraformaldehyde. After probe hybridization, Cells were pooled and loaded onto a Chromium Single-Cell Chip Q along with the reverse transcription (RT) master mix and single cell TL gel beads, aiming for 40000 cells per channel. Following generation of single-cell gel bead-in-emulsions (GEMs), reverse transcription was performed using a Applied Biosystems™ Veriti™ 96-Well Thermal Cycler (Thermo Fisher Scientific, USA). Amplified cDNA was purified using SPRIselect beads and converted to the sequencing library according to the manufacturer's protocol. Library was sequenced on an Illumina NovaSeq X platform following read length: 150 bp Read 1 (16bp single cell barcode, 10x barcode; 12bp Unique molecular identifier, UMI), 150 bp Read 2 (transcript insert), 10 bp i7 Index (sample index), 10 bp i5 Index (sample index)