Figure 5
From: Efficient generation of gene-modified pigs via injection of zygote with Cas9/sgRNA
Detection of the Npc1l1 sgRNA:Cas9-mediated off-target cleavages in vivo.
(a) PCR products of the potential off-target sites 2 and 4 (OTs 2 and 4) of Npc1l1 sgRNA:Cas9 from founder pigs. (b) Detection of Npc1l1 sgRNA:Cas9-mediated off-target cleavage at the OTs 2 and 4. All PCR products from (a) were subjected to T7EN1 cleavage assays. *, different cleavage band patterns compared to the control. The size of the digested bands does not match that expected. (c) Sequencing results of PCR products. No off-target indel mutation was observed in the T7EN7 cleavage bands. The two red arrows indicate potential off-target sites of OT2 and OT4, respectively. Con denotes the corresponding sequence of each retrieved from Ensembl.
