FIGURE 1.

Generation of CFTR^{F508del / F508del} and CFTR^{G542X / G542X} sheep fetal fibroblast cells and lambs. (A) Schematic diagram of CRISPR/Cas9 target sites in exon 11 and 12 for introduction of the F508del and G542X mutations using the ssODN_1 and 2, respectively. The blue arrows represent the primers used to amplify the exon 11 and 12. (B) Representative sequence analysis for cell colonies and lambs containing F508del, similar to the CF patients, the “CTT” nucleotides are deleted between the codons ATC and TTT. Sequencing results indicate that four isolated cell colonies and two cloned lambs contained homozygous F508del mutations. (C) Representative sequence analysis for cell colonies and lambs containing G542X mutation. Similar to the G542X CF patients, Glycine‐encoded codon “GGA” is mutated with a single nucleotide replacement, “G” to “T,” leading to the generation of stop codon, “TGA.” Sequencing results demonstrate that all nine SFF cell colonies and three cloned fetus/lambs present the same homozygous G542X mutation. The arrow represents the point mutation site and the codon sequences are framed