A mutant Bama miniature pig exhibiting the phenotype of severe CH was identified from a large-scale ENU mutagenesis screen. (A) A neonatal mutant compared with its littermate control. (B) The pedigree of the mutant family exhibiting an autosomal-recessive mode of inheritance. (C) A significant linkage peak between 120 Mb and 144 Mb on chromosome 1 was identified using a GWLS approach (LOD > 5). (D) A nucleotide A to G substitution in DUOX2 was identified by WGS and cosegregated with the mutant phenotypes. (E) The porcine DUOX2 gene is composed of 31 exons, and the mutation was located in exon 8 and resulted in a change of aspartic acid to glycine at position 409 (D409G) of the protein. (F) The D409G mutation (red arrow) is located in the peroxidase-like domain of DUOX2. The R376W and R434X mutations were found previously in patients with CH.^25,26  *, Glycosylation site. (G) The aspartic acid residue (arrow) in the D409G variant is highly conserved in mammalian DUOX2 proteins. (H) The thyroid glands of D409G pigs showed overt goiter formation at size and weight (WT, n = 7; D409G, n = 8; **P < .01). (I) Sections of the thyroid glands were stained with hematoxylin and eosin (H&E). Follicles in the thyroids of D409G pigs lost their normal shape and became convoluted as a result of highly proliferating epithelial cells (black arrows) and completely disappearing colloids (asterisks) when compared with WT littermates. Scale bars, 50 μm. (J) The serum THs of T3, T4, FT3, and FT4 produced by the thyroid glands were dramatically decreased, whereas TSH was excessively elevated, in the mutants (n = 10) compared with the WT individuals (n = 10; **P < .01). EF hand, the position of calcium binding; FAD binding, flavin adenine dinucleotide–binding motif; H, heme-binding site; Mut, mutant; NADPH binding, nicotinamide adenine dinucleotide phosphate–binding motif; TM, transmembrane domain.