Figure 1. Generation of SCN5A^E558X/+ pigs.

(A) Precordial lead ECG from the 10-year-old child with SCN5A^E555X/+ mutation, at baseline and during flecainide challenge. (B) The targeting vector contains a G-to-T point mutation (asterisk), resulting in replacement of a glutamic acid at amino acid 558 with a premature stop codon (GAG to TAG), as well as a floxed (triangles) neomycin resistance cassette (Neo^R) driven by the phosphoglycerate kinase promoter in intron 11 for selection. (C and D) Properly targeted cells were identified by Southern blot (C) and direct sequencing (D). (E) Yucatan mini pigs harboring a single copy of the SCN5A^E558X mutation (SCN5A^E558X/+ pigs; E558X/+) were viable and appeared grossly normal. (F) WBs of membrane fractions detecting Na_V1.5 and the sodium-potassium ATPase (Na/K ATPase) as loading control. Molecular weight markers (250 and 100 kDa) are indicated in lane 1. (G) Na_V1.5 protein levels (normalized to Na/K ATPase), displayed relative to WT. *P < 0.05. (H) Patch-clamp analysis of atrial myocytes. Shown are representative whole-cell Na⁺ current recordings and average current/voltage relationship from adult WT (n = 23 cells) and SCN5A^E558X/+ (n = 16 cells) left atrial myocytes. *P < 0.01.