interferon that plays an important role in both innate and cell-mediated immune responses. Because of the low similarity with their mammalian homologues, IFN-γ in teleost fish has only recently been identified. In this study, the cDNA sequence encoding the IFN-γ molecule was cloned from the orange-spotted grouper, , and its gene organization was revealed. in response to Poly I:C stimulation was also examined. The IFN-γ cDNA sequence consists of 867 bp, which encodes a putative protein of 200 amino acids. The gene is 5-kb long and contains four exons, which are comparable in size with IFN-γ genes in other vertebrates. This IFN-γ was only . Sequence analysis revealed that the IFN-γ in the orange-spotted grouper contains the typical motif and a conserved nuclear localization site in the C-terminal end and displays an alpha-helix structure similar to mammalian IFN-γ. Real time PCR analysis results showed that the expression of IFN-γ increased in the thymus, head kidney, spleen, kidney, skin, and gills following the poly I:C stimulation, indicating its role against viruses in orange-spotted grouper.