Amplicon-based NGS for NICER gene editing

We have developed a gene editing method named NICER, designed to correct heterozygous mutations. This method utilizes multiple nicks (MNs) induced by the Cas9 nickase, with the homologous chromosome serving as an endogenous repair template. To validate that NICER gene editing introduces minimal off-target mutations, we conducted the following tests:We analyzed the indel frequencies at both on-target and off-target sites for the Cas9-sgRNA complex and the Cas9(D10)-sgRNA complex using amplicon-based NGS. We examined the sequence specificity of the Cas9-sgRNA complex using amplicon-based NGS.