The usage and value of soybean [('Glycine max' (L.) Merrill] oil is affected by its fatty acid profile. The objectives of this study were to: (1) characterize the inheritance of the altered fatty acid levels in the soybean lines developed at the University of Guelph; (2) investigate the environmental effects on the performance of RILs developed from crosses among the se lines and analyze the relationships among fatty acid contents, seed quality and agronomic traits; (3) map QTLs for altered fatty acid levels and investigate QTLxenvironment interactions. The inheritance study indicated that palmitic acid segregation involved two loci, 'Fap1' had an additive genetic effect and 'Fapx' exhibited partial dominance. Stearic acid segregated at the 'Fas' locus with a dominant genetic effect. Linolenic acid segregated at the 'Fan' locus with an additive genetic effect. Combined analysis of variance indicated that entry * environment interaction effects were significant for all traits. Year effects were larger than location effects for palmitate, stearate, and oleate contents, whereas location effects were greater than year effects for all other traits including linoleate, linolenate, protein, and oil contents, plant height, lodging, maturity and yield. The climatic data indicated that the minimum daily temperature rather than the mean and maximum was the most important factor affecting the unsaturated fatty acid levels. Trait correlations varied upon the population, generations and growing environment. There was a general lack of correlations between the fatty acid composition and agronomic traits. Both the 'Fas' and 'Fan' loci were mapped to LG B2 at a 21 cM distance in between. Markers located at 'Fas' and 'Fan' locus explained over 70% and 80%, respectively, of the phenotypic variation for stearic acid and linolenic acid levels. The elevated stearic acid level conferred by 'Fas' locus was most likely to be the result of reduced activity of stearoyl-ACP desaturase which catalyzes the reaction form stearic acid to oleic acid in the fatty acid biosynthesis pathway. Molecular markers with high R2 values, or the QTLs with high LOD scores were more likely to be detected in multiple environments. No consistent segregation distortion pattern for markers was detected between the two mapping populations used in this study.