GSM7635028: Tumor cells enriched from mouse Undifferentiated Pleiomorphic Sarcoma tumors (p53-/-Ccne1+) (tech rep 1); Mus musculus; RNA-Seq
Sarcoma tumors were digested into single-cell suspensions using the Mouse Tumor Dissociation Kit and gentleMACS dissociator (Miltenyi) and filtered twice through 70um filters before ACK red blood cell lysis. Cells were washed in FBS-containing medium, stained with antibodies, and enriched by MACS or FACS. Where applicable, each individual biological replicate (mouse tumors) was stained with a unique hashtag oligonucleotide antibody (Biolegend TotalSeq-A). Biolgeical replicates were then combined in equal proportions into a single pool of cells prior to capture. For each library, 16,000 cells were loaded into one well of a 10X Genomics Chromium chip for NextGEM 3' v3.1 droplet sequencing, at a concentration between 600-1400 cells/uL, thus targeting capture of 10,000 cells per sample. Each 10X NextGEM 3' v3.1 library was constructed according to manufacturer protocols and sequenced on the NovaSeq 6000, targeting 20,000 paired-end reads per cell for gene expression libraries, or 500 reads per cell for hashtag libraries.