Phylogenetic analysis demonstrates that vertebrate sodium channel β1-4 subunit genes are orthologous, that β1/β3 and β2/β4 are closely related, and that zβ4.1 and zβ4.2 resulted from a recent gene duplication in fish. Actual (human, zebrafish) and predicted (chicken, frog) amino acid sequences of β subunit and related genes were aligned using CLUSTALX (v1.83). Phylogenetic trees were reconstructed using the neighbor-joining method of Saitou and Nei and viewed with NJPlot software. Alignment gaps were excluded and the Kimura correction was made for multiple substitutions. Bootstrapping (n = 1000) was applied to test the robustness of each node. Tree is unrooted due to the lack of evidence for β subunit-like genes in invertebrate species. HUGO gene nomenclature symbol IDs: MPZ = myelin protein zero; MPZL1 = myelin protein zero-like gene isoform A; EVA1 = epithelial V-like antigen 1; EVA1L = unannotated gene similar to EVA1.