Camellia is an ornamental tree that is grown worldwide. There are many excellent classical cultivars whose origins are unclear. Chloroplast DNA (cpDNA) is a good material for tracing maternal lineages and we therefore conducted polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the cpDNA atpI-atpH region from 162 accessions derived from 49 species of the genus Camellia. In order to specifically and accurately amplify the atpI-atpH region, camellia-specific primers were designed and utilized with some of the accessions. PCR amplification of the atpI-atpH region resulted in two major bands of approximately 800 bp and 1200 bp. TaqI digestion resulted in 3 different band patterns derived from the 800 bp fragment and 5 patterns derived from the 1200 bp fragment. Most of these eight PCR-RFLP patterns were distributed widely amongst species and sections of the genus. In particular, it was possible to use this approach to distinguish between the main species from which ornamental camellia cultivars are derived. Sequence analysis of the atpI-atpH region showed a number of mutations, which were characteristic of particular species. These results indicate that PCR-RFLP analysis of the atpI-atpH region will be very useful for investigating variations in the genus Camellia.