GO-CRISPR: a highly controlled workflow to discover gene essentiality in loss-of-function screens

SUMMARY Genome-wide CRISPR screens are an effective discovery tool for genes that underlie diverse cellular mechanisms that can be scored through cell fitness. Loss-of-function screens are particularly challenging compared to gain-of-function because of the limited dynamic range of decreased sgRNA sequence detection. Here we describe G uide- O nly control CRISPR (GO-CRISPR), an improved loss-of-function screening workflow, and its companion software package, T oolset for the R anked A nalysis of GO- C RISPR S creens (TRACS). We demonstrate a typical GO-CRISPR workflow in a non-proliferative 3D spheroid model of dormant high grade serous ovarian cancer and demonstrate superior performance to standard screening methods. The unique integration of the pooled sgRNA library quality and guide-only controls allows TRACS to identify novel molecular pathways that were previously unidentified in tumor dormancy. Together, GO-CRISPR and TRACS can robustly improve the discovery of essential genes in challenging biological scenarios.