Abstract: The complete coding sequence of glutathione peroxidase 1 (GPx1) of Nile tilapia (Oreochromis niloticus) was cloned by rapid amplification of cDNA ends (RACE).The GPx1 contains 984 bp, including a 56 bp 5′-untranslated region, a 576 bp coding sequence (CDS), a 352 bp 3′-untranslated region and a 20 bp PolyA tail. A selenocysteine (Sec) was encoded by the unusual stop codon, TGA, with the selenocysteine insertion sequence (SECIS) located at 3′UTR. The GPx1 was predicted to encode 191 amino acids, and its molecular weight was 21.8 kDa with a pI of 8.04; neither signal peptide nor N-Glycosylation site was found. The analysis showed that GPx1 was a non-transmembrane protein, and it possessed classic catalytic tetrad comprising selenocysteine, tryptophan, glutamine and asparagine. We compared GPx1 between Nile tilapia and other vertebrate animals, and found that the similarity of nucleotide acid sequence was 43.2%~58.2%, and that of amino acid sequence was 58.1%~80.6%. Phylogenic analysis indicated that GPx1s of different classes of vertebrates split into different clusters. The 3D structure of tilapia GPx1 was predicted with Swiss-Model software, and sequence analysis suggested that GPx1 was homotetrameric.