In contrast to animal mitochondrial DNA, intraspecific variation in chloroplast DNA is thought to be very rare. This presumption has prevented the application to plant population biology of the diversity of molecular genetic techniques now well established for animal mitochondrial DNA. In Trifolium pratense, however, extensive intrapopulational variation does exist. In two paper I report a characterization of unprecedented restriction fragment profile variation within single populations. Populations typically contain a common genotype and many rare ones; often the rare genotypes differ from population to population. While both nucleon and nucleotide diversity, as well as estimates of Wright's F(ST), indicate a large within-population component and relatively little diversity among populations, the distribution of plastid genotype frequencies in each population is not homogeneous. Estimates of migration rate based on chloroplast DNA genotypes suggest a moderate number of migrants per generation. The unusually high level of genetic variation in Trifolium chloroplast DNA provides the first opportunity to use the plastid genome of plants to study population differentiation. Furthermore, it suggests that the plastid genome may not be as invariant as previously believed, but may instead exhibit high levels of genetic diversity at the population level.