Genome-wide CRISPR screen in standard growth conditions and stem cell conditions using primary low-passage KPf/fC pancreatic cancer cell lines

Purpose: The goal of this study is to identify novel genes required for the survival and proliferation of pancreatic cancer cells in standard culture conditions and in stem cell conditions Methods: Three independent primary pancreatic cancer cell lines were established from endstage Musashi2-reporter KPf/fC mice, and transduced with the genome-wide lentiCRISPR v2 library. Cells were grown under puromycin selection for 3 weeks (6 passages), then plated in non-adherent stem cell conditions in single-cell density for 1 week. Genomic DNA was isolated directly after library transduction, after selection in standard growth conditions, and after sphere growth in stem cell conditions. For sequencing, guide RNA's were amplified from total genomic DNA and barcoded for sequencing on an Illumina HiSeq2500 using V4 sequencing chemistry Conclusions: Our study represents the first genome-wide CRISPR screen identifying genetic vulnerabilities of KPf/fC cells in stem cell conditions using 3 biologically unique primary cell lines. Overall design: A multiplexed genome-wide CRISPR dropout screen was performed using primary KPf/fC cell lines in standard growth conditions and in stem cell conditions.