Yield of Pima cotton (Gossypium barbadense L.) has tripled over the last forty years with the development of new cultivars and strains. Six genetic lines representing successive stages in the breeding process (one primitive non-cultivated accession, four cultivars with release dates from 1949 to 1983, and one unreleased breeding line) were grown in a greenhouse, and their diurnal gas exchange properties were compared. Among the cultivated types, genetic advances were closely associated with increasing single-leaf photosynthetic rate (A) and stomatal conductance (g(s)), especially in the morning. In midafternoon, A and g(s) of these lines tended to converge. The primitive line behaved differently, with A and g(s) near those of the cultivated types early in the morning, but much lower than them for the rest of the day. In both morning and afternoon, A was correlated with g(s) but not with leaf thickness, concentrations of chlorophyll or starch, or intercellular CO₂ concentration (c(i)). In the oldest cultivar, the relationship of A to c(i) changed only slightly from morning to afternoon. In the two most recent lines, the slopes of the A:c. curves at limiting c(i) exceeded those of the oldest cultivar by 25 to 50% in the morning, but the differences were much smaller in the afternoon. The maximum A of the newer lines at high c(i) exceeded that of the oldest cultivar only in the morning. Breeding for increasing yield has enhanced the photosynthetic capacity and stomatal conductance of Pima cotton, and altered the diurnal regulation of photosynthesis. Nonetheless, no evidence indicates that increased photosynthetic rate is directly responsible for increased yield.