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The present study investigates the effect of different levels of galactooligosaccharide (GOS) on innate immune parameters, immune related genes expression as well as growth performance in zebrafish (Danio rerio). Four hundred and twenty fish (mean weight 45±0.1mg) were supplied, randomly stocked in twelve aquaria assigned to four treatments. Zebrafish were fed with either control diet or control diet enriched with different levels (0.5, 1 and 2%) of GOS for eight weeks. At the end of feeding trial innate immune parameters (total immunoglobulin, total protein and alkaline phosphatase activity), immune related genes expression (interleukin 1 beta [il1b], Lysozyme [lyz], tumor necrosis factor alpha [tnf-alpha]) as well as growth performance were measured. Evaluation of immune parameters revealed significant (P<.05) increase of total protein and total Ig in zebrafish fed 1 or 2% GOS compared other treatments. However, in case of lysozyme activity no significant (P>.05) differences were noticed between GOS fed fish and control group. Also, in case of ALP activity, significant increase (P<.05) was observed in 2% GOS treatment. Gene expression studies revealed significant upregulation (P<.05) of tnf-alpha and lyz genes in GOS fed fish. While no significant (P>.05) difference was observed in case of il1b gene expression. Evaluation of growth performance at the end of feeding trial revealed no significant (P<.05) improvement in GOS fed groups. The present results revealed positive effects of GOS on innate immune parameters and related gene expression in zebrafish.Galactooligosaccharide [GOS] improved innate immune parameters in Zebrafish.GOS upregulated the expression of immune related genes in Zebrafish.GOS had no remarkable effects on growth performance of Zebrafish.