Comparative Molecular Analyses of LNCaP and Substrains Identify Drivers of Cancer Behavior and Therapy Resistance [scRNA-seq]

Comparative Molecular Analyses of LNCaP and Substrains Identify Drivers of Cancer Be-havior and Therapy Resistance To collect single cell suspension, cells were trypsinized and washed once in PBS and re-suspended in cell culture media followed by hemocytometer cell count to ensure cell viability above 70%. The cell suspension were loaded into Chromium Next GEM Chip G (10x Genomics , PN#1000120) with 3′ v.3.1 beads (10x Genomics , PN#2000164) and partitioning oil. Individual cells were barcoded using a 10× Chromium Controller (10x Genomics). RNA from the barcoded cells were subsequently reverse-transcribed and amplified with a Chromium Single Cell 3′ v3.1 Reagent Kit (10x Genomics, PN#1000123). Libraries were subsequently constructed from 25% of amplified cDNA traces from pre-vious step with reagents from a Chromium Single Cell 3′ v3.1 Library Kit (10x Genomics, PN#1000190). The cDNA and library traces were quantitated and identified by Tapestation. Sequencing were performed with an Illumina NextSeq 2000 P3. All cells grown in RPMI1640 supplemented with 10%FBS and 1%Pen/Strep antibiotic.