A characteristic feature of grasses and commercially important cereals is the presence of (1,3;1,4)-beta-d-glucans in their cell walls. We have used comparative genomics to link a major quantitative trait locus for (1,3;1,4)-beta-d-glucan content in barley grain to a cluster of cellulose synthase-like CslF genes in rice. After insertion of rice CslF genes into Arabidopsis, we detected (1,3;1,4)-beta-d-glucan in walls of transgenic plants using specific monoclonal antibodies and enzymatic analysis. Because wild-type Arabidopsis does not contain CslF genes or have (1,3;1,4)-beta-d-glucans in its walls, these experiments provide direct, gain-of-function evidence for the participation of rice CslF genes in (1,3;1,4)-beta-d-glucan biosynthesis.