Zymograms of glucosephosphate isomerase (GPI) and phosphogluconate dehydrogenase (PGD) revealed three isozymes for each enzyme in the plant species Scabiosa columbaria. Intergenic heterodimers are formed between the polypeptides coded by Gpi-1 and Gpi-2 and between those coded by Pgd-1 and Pgd-2, indicating that a GPI and a PGD locus have been duplicated in the past. The ancestral genes assort independently with their duplicated gene, suggesting that the duplications have originated from a process of translocation. Linkage was found only between Gpi-1 and Pgd-2 and between Gpi-2 and Pgd-1, suggesting that the duplicated loci were located on the same translocated chromosomal segment. Both duplications are present in all other examined species of Scabiosa and in Cephalaria and Knautia, two other genera of the Dipsacaceae. The genera Succisa and Dipsacus, also belonging to the Dipsacaceae, do not show Gpi-1 activity, making Gpi-2 and Pgd-1 the most likely ancestral genes. In Succisa, the isozymes of Gpi-1 and Gpi-2 either overlap or Gpi-1 has been silenced. The combined results suggest that a chromosomal segment containing Gpi-2 and Pgd-1 has been translocated before the divergence of Scabiosa, Cephalaria, Knautia, and Succisa.