title: PyMT-N tumor-infiltrating immune cells
design: Tumors were processed into single-cell suspensions. To enrich tumor-infiltrating lymphocytes, CD11b+ cells were depleted through magnetic sorting, followed by FACS sorting of CD45+ cells for 3' scRNA-seq. The single-cell gene expression Library was prepared according to the Chromium Single Cell Gene Expression 3v3.1 kit (10x Genomics). In brief, single cells, reverse transcription (RT) reagents, Gel Beads containing barcoded oligonucleotides, and oil were loaded on the Chromium controller (10x Genomics) to generate single-cell GEMS (Gel Beads-In-Emulsions) where full-length cDNA was synthesized and barcoded for each single cell. Subsequently, the GEMS are broken and cDNA from each single cell is pooled. Following cleanup using Dynabeads MyOne Silane Beads, cDNA is amplified by PCR. The amplified product is fragmented to optimal size before end-repair, A-tailing, and adaptor ligation. The final library was generated by amplification. The libraries that passed the quality control, were sequenced by the NovaSeq 6000 (Illumina).