A key question is whether a gRNA position capable of activating transcript expression via dCas9−VP64 or dCas9−VPR will also reliably repress transcription for the same gene via dCas9−KRAB. If true, this would reduce the necessary design, synthesis, cloning, and validation of different gRNAs for each experiment, facilitating more scalable experimental execution. We used the same gRNA positions examined in the activation setting (Figure 2), but in NPCs and neurons expressing dCas9−KRAB fusion proteins (Figure 3). Again, both antibiotic-selected (Figure S2) and non-selected lentiviral-transduced dCas9−KRAB NPCs were evaluated. Overall, gRNA efficacies with dCas9−KRAB (Figure 3) were only sometimes consistent with those achieved for dCas9−VP64 or dCas9−VPR (Figure 2).
