Interferon gamma (IFN-gamma) has important roles in both innate and adaptive immune responses. In this study, the cDNA and genomic sequences of Atlantic cod IFN-gamma were cloned and found to encode a putative protein containing 194 amino acids with a 24 amino acid signal peptide sequence. The gene is composed of four exons and three introns similar to IFN-gamma genes of other vertebrates. The cod IFN-gamma showed only 14-29% amino acid identity with other fish IFN-gamma and 9-17% identity with IFN-gamma from higher vertebrates. However, cod IFN-gamma possesses the typical IFN-gamma motifs in the C-terminal end of the protein and displays an alpha-helix structure similar to mammalian IFN-gamma. The promoter region contains a putative ISRE element indicating up-regulation by type I IFNs and dsRNA. Real time RT-PCR analysis confirmed that IFN-gamma gene expression was up-regulated in organs of cod injected with the dsRNA polyinosinic:polycytidylic acid (poly I:C), which is a strong inducer of type I IFNs. Injection of cod with formalin-killed Vibrio anguillarum also increased IFN-gamma expression in head kidney, but to a much lesser extent than poly I:C. The gene expression results thus indicate a role for IFN-gamma in innate immune response against both virus and bacteria in Atlantic cod.