Figure 1.

Design and execution of a CRISPR/Cas9 tiling screen. (A) Schematic of a CRISPR–Cas9 tiling screen and phenotypic readout. The percent of population containing each genotype is calculated based on in-frame repairs occurring at 20% of edits (Chakrabarti et al. 2019), and a hypothetical example of how this reveals functional protein motifs is shown below. (B) Cartoon representation of key molecular activities of kinetochore- and microtubule-mediated processes. Genes/proteins to be screened are listed at the right according to their best-characterized function. (C) Characteristics of a tiling library targeting mitotic factors. (D) Schematic of a proliferation-based screening approach using lentiviral particles to insert both spCas9 and sgRNA sequences into genomic DNA. This allows next-generation sequencing of sgRNA sequences to serve as an indirect readout of their effect on cell growth.