Fig. 1. Design of CRISPR-Cas9 targeting of porcine PKD1 and isolation of nuclear donor cells.

A Schematic representation of CRISPR-Cas9 targeting site for porcine PKD1 gene. The gRNA targeting sequences are underlined. The black box represents a protospacer adjacent motif (PAM) sequence. B CRISPR-Cas9-induced mutations in the isolated cell clones. The deletion mutation is indicated by asterisks. C Direct DNA sequence analysis of a PKD1^insG/+ nuclear donor cell (clone #132). D Deduced amino acid sequence resulting from the induced mutation (PKD1^insG) in cell clone #132. The amino acid sequences in red indicate nonsense amino acids. A 1-bp insertion of guanine nucleotide in porcine PKD1 created a BsrI restriction enzyme site (underlined) and stop codon at amino acid residue 113.