Reproductive proteins typically have high rates of molecular evolution, and are assumed to be under positive selection from sperm competition and cryptic female choice. However, ascribing evolutionary divergence in the genome to these processes of sexual selection from patterns of association alone is problematic. Here, we use an experimental manipulation of postmating sexual selection acting on populations of house mice and explore its consequences for the expression of seminal vesicle secreted (SVS) proteins. Following 25 generations of selection, males from populations subjected to postmating sexual selection had evolved increased expression of at least two SVS genes that exhibit the signature of positive selection at the molecular level, SVS1 and SVS2. These proteins contribute to mating plug formation and sperm survival in the female reproductive tract. Our data thereby support the view that sexual selection is responsible for the evolution of these seminal fluid proteins.