Pheromone-binding proteins (PBPs) are soluble transporter proteins that increase the capture and the solubilization of pheromone molecules in the lymph surrounding the olfactory receptors. A polymerase chain reaction-based method was used to identify PBP genes in Agrotis species for an evolutionary genomic study of noctuid moth PBPs. From genomic DNA we determined the structure of different PBP genes in the two closely related species, Agrotis ipsilon and A. segetum. In all, we clearly identified four genes (Aips-1, Aips-2, Aseg-1 and Aseg-2) that represent two distinct PBP orthology groups. We found that the four genes have the same exon-intron structure and that they comprise three exons and two introns but differ in length mainly in the second intron. The three exons of Aseg-2 and Aips-2 have the same lengths but both intron 1 and intron 2 differ in length between the genes. In contrast, Aips-1 and Aseg-1 show dissimilarity only in the length of intron 2. Interestingly, introns 1 and 2 are inserted in the same positions in the Aips-1, Aips-2, Aseg-1 and Aseg-2 genes. These findings show that the Agrotis PBP genes have common ancestry and probably originate from gene duplication before the speciation of ipsilon and segetum. We found that expression of Aips-1/Aseg-1 and Aips-2/Aseg-2 is antennal-specific, but expression is not restricted to the male antennae.