An important aspect of the safety assessment of genetically modified crops to be used for human food and animal feed is the product composition, including nutrients and antinutrients. Cotton lines have been developed that are tolerant to glyphosate, and the active ingredient in the herbicide Roundup. The glyphosate-tolerant lines were generated by the stable insertion of a glyphosate tolerance gene in cotton cv. Coker 312. The glyphosate tolerance gene encodes for 5-enolpyruvylshikimate-3-phosphate synthase (3-phosphoshikimate 1-carboxyvinyltransferase) (EPSPS) from Agrobacterium strain CP4 (CP4 EPSPS). The composition of the cottonseed and oil from two glyphosate-tolerant lines, 1445 and 1698, was compared to that of the untransformed Coker 312 and to published values for other commercial cotton varieties. The nutrients measured were protein, fat, fibre, carbohydrate, calories, moisture, ash, amino acids, and fatty acids. The antinutrients measured included gossypol, cyclopropenoid fatty acids, and aflatoxins. In addition, the fatty acid profile and alpha -tocopherol levels were measured in the refined oil. These analyses demonstrated that the glyphosate-tolerant cotton lines are compositionally equivalent to the parental and conventional cotton varieties commercially available.