Figure 1: Analysis of RGN-induced genomic mutations in NIH/3T3 cells.
From: Efficient generation of FVII gene knockout mice using CRISPR/Cas9 nuclease and truncated guided RNAs
(A) PCR amplicons of target sites in the FVII in cells transfected with RGN expression plasmids (F7–1, F7–2, F7–3, tF7–1, tF7–2, and tF7–3) were subjected to T7EI assays. In each RGN group, T7EI-digested samples (a) and untreated controls (b) were shown. Samples of wild type (WT) cells served as negative controls. (B) Off-target cleavage induced by RGNs of F7–1 (a) and tF7–1 (b) at five potential off-target sites (OT1–1, OT1–2, OT1–3, OT1–4, and OT1–5) in transfected cells were detected. Samples from WT cells served as controls (c). M, marker. The fragments of T7EI digested mutant type (MT) were indicated out by arrow. The target and off-target sites were subsequently confirmed by PCR-sequencing.
