Description: Delivery of CRISPR/Cas9 via bioreducible lipid nanoparticles (LNPs) to the inner ear in Ai14 mice. Subset of mice were administered LNP via canalostomy injection compared to uninjected control mice. Tissues were harvested 6 days after LNP administeration. On-target and off-target editing was assessed.

Editing Assay: On-target editing analyzed by confocal imaging of whole mouse cochlea for direct fluorescence RFP (tdTomato) compared to Hoeschst stained nuclei. Off-target editing analysis: peripheral tissues were fixed, cryosectioned and imaged directly for RFP (tdTomato). Presence or absence of signal above background was recorded for each tissue assessed. For 306-O12B, On-target N=6; Off-target N=5 (2 males, 3 females); for 306-S10, On-target N=6; Off-target N=1 (female); and for Uninjected Control, On-target and Off-target N=2.

Parent Project: The Jackson Laboratory Gene Editing Testing Center (JAX-GETC) - SATC-Chen, Submission 2 (On- and Off-target data)

• Other experiments in this project: 1

  • Repeat experiment of independent validation of Chen delivery platform using LNPs to deliver CRISPR/Cas9 to mouse inner ear

Original Experiment/s that are being validated:

• Testing new LNPs (lipid nanoparticles) for delivery of Cas9 mRNA/sgRNA in adult mouse cochlea