Workflow of pooled vs arrayed CRISPR-Cas9 screening approaches. For either screening strategy, the input pool of sgRNAs and Cas9 is designed using complex bioinformatic algorithms to maximize targeting efficiency and limit off-target effects. In the pooled screening approach, a viral sgRNA library generated from the sgRNA/Cas9 complexes can then be delivered to a single vessel of cells at a low MOI. In contrast, within the arrayed screening format viral libraries are delivered to discrete cell populations grown in multiwell plates to allow for unique representation of individual sgRNAs within each well. Following the administration of a challenge (such as drug treatment), based on the selection strategy adopted (positive, negative, or FACS-associated marker based), several rounds of cell selection may be included within pooled screens to allow for targeted cell enrichment. In contrast, the arrayed screens do not require selective enrichment, rather the developing phenotype within the individual cell populations form the raw data that can be visualized in the final step of the assay through high-throughput image acquisition strategies followed by statistical ranking of the observed phenotypes. In contrast, raw data (genomic DNA from the selected cell populations) must be profiled using deep-sequencing approaches followed by an analysis of depleted or enriched sgRNA population to identify the potential genes that could be associated with potential cancer pathways although downstream validation will be required to confirm the speculative role of the potential genes within the disease. Key steps of convergence between the 2 pathways have been highlighted for both pathways. gRNA, guide RNA. Created with www.BioRender.com.