FIG 1.

Toxoplasma genome-wide loss-of-function screen in naive or IFNγ-activated HFFs. (A) Screening procedure. RH-Cas9 parasites were transfected with linearized plasmids containing 10 sgRNAs against every Toxoplasma gene. Transfected parasites were passaged for 4 to 5 passages in HFFs under pyrimethamine selection to remove nontransfected parasites and parasites that integrated plasmids with sgRNAs targeting parasite genes important for fitness in HFFs. Subsequently, the pool of mutant parasites was passaged 4 times in naive or IFNγ-stimulated HFFs. (B) The sgRNA abundance at different passages, determined by Illumina sequencing, was used for calculating phenotype scores and identifying genes that confer fitness specifically in IFNγ-activated HFFs. Genes shown in Table 1 are indicated with a red dot.