title: GSM8598050: corin_PD1_2; Mus musculus; RNA-Seq
library_construction_protocol: Minced tumor biopsies were incubated with 200uL 20mg/mL collagenase, 200uL 20mg/mL hyaluronidase, and 5uL DNase for 20 minutes to 1 hour with agitation until the tumor fully dissociated. Single-cell suspension was passed through a 100um strainer and washed with 1X PBS by centrifugation. Cells were resuspended in 1ml of freezing media frozen down. Cryopreserved single-suspension tumor samples were thawed and dead cells were removed using the Dead Cell Removal Kit (Miltenyi, 130-090-101) following manufacturer instructions. Live cells from the flow-though were used for CD45+ cell isolation using CD45 mouse MicroBeads (Miltenyi, 130-052-301) following the recommendations. CD45+ cells were counted and resuspended in PBS + 0.04% BSA at 1000 cells/µL for scRNA-seq. About 23,000 cells were loaded onto a 10× Genomics ChromiumTM X instrument (10x Genomics) according to the manufacturer's recommendations. The scRNAseq libraries were processed using Chromium GEM-X Single Cell 5' Kit v3 (10x Genomics). Quality controls for amplified cDNA libraries and final sequencing libraries were performed using Bioanalyzer High Sensitivity DNA Kit (Agilent). The sequencing libraries for scRNAseq were normalized to 4nM concentration and pooled. The pooled sequencing libraries were sequenced on the Illumina NovaSeq S4 300 cycle platform. The sequencing parameters were: Read 1 of 28bp, Read 2 of 90bp, Index 1 of 10bp and Index 2 of 10bp.