The quantitative genetic basis of fatty acid composition was examined in the Genetically Improved Farmed Tilapia (GIFT) strain of Nile tilapia selected for high breeding value for body weight and in the contemporaneous control selected for average breeding value. Gas chromatography analysis of 514 frozen fillet samples, obtained from the offspring of 104 sires and 154 dams from two generations in 2006 and 2007, showed that the fish possess all important fatty acids (FA), with the amount of omega-3 polyunsaturated fatty acids being 3.6%. The ratio of unsaturated to saturated fatty acids was greater than one (1.4) in the GIFT fillets, which is in line with the current dietary recommendation. Genetic analysis of FA was combined with 5532 performance and fillet records, and 2000 fillet samples analyzed for flesh composition and flesh quality attributes. A restricted maximum likelihood method applied to bivariate or trivariate mixed model was carried out in a full pedigree comprising a total of 18,970 animals to estimate genetic parameters and correlated responses in FA composition to the selection program. The estimates of heritability for individual FA varied from zero to medium, suggesting that FA composition of the GIFT fillet can respond to selection. Genetic correlations among FA within the same group (saturated, monounsaturated or polyunsaturated FA) were generally synergetic, but were antagonistic between groups. There were also both favourable and unfavourable genetic associations between important high chain polyunsaturated fatty acids and performance, fillet and flesh quality traits. Correlated changes in FA composition were measured by comparing least square means of the selection and control lines. Overall, selection for high growth resulted in very little change in FA composition. One notable change was that the content of heptadecanoic acid (C17:0) decreased in the selection relative to control line, which is beneficial for human health. In summary, the set of genetic parameters estimated and the response realized in the GIFT strain indicate that FA composition of tilapia or fish in general can be altered by genetic means.