Multiplexed CRISPR-Cas9 system in a single adeno-associated virus to simultaneously knock out redundant clock genes

Key findings

First demonstration of triple sgRNA multiplexing in single AAV for simultaneous knockout of gene families. Used GoldenGate assembly to create U6-U6-U6 cassettes with hSyn-mCherry-KASH reporter. Successfully targeted circadian clock genes (Crys, Pers, Bmal1) with efficient editing in vitro and in vivo.

Canonical designs

Parts used