A panel of AsCas12a variants harboring combinations of mutations are tested using crCD55-4 and crCD81-1 using the fusion protein domain architecture shown. Both AsCas12a fusion protein and crRNA constructs are delivered by lentiviral transduction. D908A is a mutation in the RuvC catalytic triad that renders Cas12a DNase inactive 24 , 54 . Other mutations are described in detail in the main text. Shown are single-cell distributions of target gene expression assayed by flow cytometry 6 days after crRNA transduction for one of three independent replicates. Additional replicates and results for additional crRNA constructs (up to 3-plex crRNA constructs) are summarized in Supplementary Fig. 6a–c.
