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Restriction fragment patterns of mitochondrial DNA from sibling species of cyst nematodes Heterodera glycines and H. schachtii were examined. Fourteen restriction endonucleases recognizing four, five, and six base-pair sequences yielded a total of 90 scorable fragments of which 10% were shared by both species. Mitochondrial genome sizes for H. glycines and H. schachtii were estimated to be 22.5-23.5 kb and 23.0 kb, respectively. A single wild type mitochondrial genome was identified in all populations of H. glycines examined, although other mitochondrial genomes were present in some populations. The H. schachtii genome exhibited 57 scorable fragments, compared with 33 identified in the H. glycines wild type genome. The estimated nucleotide sequence divergence between the two species was p = 0.145. This estimate suggests these species diverged from a common ancestor 7.3-14.8 million years ago.