Studying and improving the IscB-ωRNA system for targeted genome editing

Enhanced OgeuIscB shows a 30.4-fold improvement in insertion-deletion formation efficiency in human cells.

• OgeuIscB with eight amino acid substitutions significantly increased DNA-binding affinity by fourfold.
• The engineered OgeuIscB-ωRNA systems achieved up to 87.3% indel and 62.2% base-editing frequencies across 26 target sites.
• Both wild-type and enhanced OgeuIscB-ωRNA exhibited moderate fidelity in genome editing.
• Off-target analysis identified key factors affecting target selection, including specific motifs adjacent to the target site.

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