Fig. 5: No detectable genome-wide off-targeting by enFnCas9 variants in human cells.
From: PAM-flexible Engineered FnCas9 variants for robust and ultra-precise genome editing and diagnostics
a Circos plot showing the comparative off-targeting profile by SpCas9 and SpRY programmed with 20-nt spacer containing sgRNA, and en1, en15, and en31 programmed with 21-nt spacer containing sgRNA against HBB locus in HEK293T cells, as captured by Digenome-seq. The scatter dots indicate the on-target site (marked by arrowhead) and off-target sites (n = 1). Each concentric circle corresponds to each Cas9 variant, as labeled in the figure. b Bar plot showing the number of off-target sites plotted on the Y-axis from Digenome-seq assay for SpCas9, SpRY, en1, en15, and en31 at HBB locus in HEK293T cells. c UpSet plot showing the hits identified in the Digenome-seq at HBB locus across Cas9 variants. The on-target is outlined with a dotted red rectangle. d Bar plot showing the indel events (%) at HBB locus by en1 plotted on the Y-axis as obtained from amplicon sequencing of the Digenome-seq detected off-targets (OTs) in HEK293T cells as compared to the on-target (ON) (n = 1). The mismatches of the off-targets are shown in lowercase and highlighted in red. OT6 was a drop-out due to PCR failure. Source data are provided as a Source Data file.
