GSM9258335: scRNA-seq of tumor-infiltrating neutrophils from trametinib-treated mice; Mus musculus; RNA-Seq
Neutrophils were isolated from tumors by fluorescence-activated cell sorting (FACS). Cells were concentrated to 1,000 cells/μL in 1× PBS with 1% BSA and loaded into the Chromium Controller (10x Genomics) with a target recovery of ~8,000 cells per sample. Cells were partitioned with barcoded gel beads and reagents from the Chromium Next GEM Single Cell 3′ v3.1 chemistry (10x Genomics). Following reverse transcription, cDNA was amplified, and gene expression libraries were constructed according to the Chromium Next GEM Single Cell 3′ v3.1 protocol (10x Genomics). Single-cell libraries were pooled and sequenced on an Illumina NextSeq 2000 P1 flowcell using a non-symmetric 28-cycle + 90-cycle paired-end configuration. All samples yielded >9 million reads per sample. Sequencing quality was high, with >93.9% of bases in the barcode regions, >92.2% of bases in the reads, and >95.6% of bases in the UMI regions reaching Q30 or above. Demultiplexing was performed allowing one mismatch in barcodes. Raw data were processed with Cell Ranger v8.0.1 using default parameters, with adjusted cell calling thresholds applied for neutrophils.