Mitochondrial DNA, widely applied in studies of population differentiation in animals, is rarely used in plants because of its slow rate of sequence evolution and its complex genomic organization. We demonstrate the utility of two polymorphic mitochondrial tandem repeats located in the second intron of the nad1 gene of Norway spruce. Most of the size variants showed pronounced population differentiation and a distinct geographical distribution. A GenBank search revealed that mitochondrial tandem repeats occur in a broad range of plant species and may serve as a novel molecular marker for unravelling population processes in plants.