FIG 1.

A high-throughput screen for AAV restriction factors reveals Crumbs 3 (Crb3) as a lead candidate. (A) Schematic of iterative FACS sorting of a Huh7 GeCKO library. The library was transduced with rAAV9-scGFP and sorted to isolate highly GFP-positive cells, which were then grown out and sorted again to separate out stably GFP-positive cells from those that became GFP negative over time. GFP-positive cells were subjected to a second sort with rAAV9-ssRFP, while GFP-negative cells were sorted again with rAAV9-scGFP. (B) Guide enrichment across iteratively sorted cells, where bubbles represent guides populations, bubble size represents relative enrichment, and bubbles highlighted in orange represent Crb3. (C) Guide representation following first- and second-round sort populations, where the proportion of the pie chart represents the proportion of reads mapping to guides for the indicated genes.