Volcano plot illustrating the differential expression of sgRNAs (shown as the name of the targeting genes). (A) Sequencing results from the one-round screen. The fold change indicates the enrichment of these sgRNAs in the lowest 30% of the GFP/mCherry cells compared with the highest 30% of the GFP/mCherry cells. The top candidate genes are highlighted. (B) Sequencing results from the two-round screen. The fold changes are calculated from the sgRNAs in the sorted cells compared with unsorted cells. (C) Flow cytometry analysis of cells expressing Ub–GFP–SKL using two different sgRNAs for each candidate gene knockout.