Gene duplication plays an important role in the evolution of organisms by allowing functional innovation and the divergence of duplicate genes.Previous studies found two Pl-like genes in grass species,suggesting functional divergence between the paralogous copies.Here,we reconstructed the evolutionary history of two PI genes from major lineages of grasses and other monocot species,and demonstrated that two PI genes (PI1 and PI2) arose from a whole genome duplication that occurred in a common ancestor of extant grasses.Molecular evolutionary analyses at the family and tribal levels found strong purifying selection acting on two genes in grasses,consistent with the conserved class B function of the PI genes.Importantly,we detected different patterns of selective relaxation between the duplicated PI genes although no signature of positive selection was found.Likelihood ratio tests revealed that the ω ratio for M domain is significantly higher in PI1 than in PI2 but that for K domain is significantly higher in PI2 than in PI1.These findings imply that complementary selective relaxation occurs in two PI genes after duplication,and provide additional molecular evidence for the subfunctionalization of the duplicated PI genes in grasses.