The psbA gene, encoding the D1 protein of photosystem II, was found to be duplicated in the chloroplast genome of two pine species, Pinus contorta and P. banksiana. Analysis of cloned overlapping restriction fragments of P. contorta chloroplast DNA showed that the two psbA genes have the same orientation and are separated by approximately 3.3 kb. The nucleotide sequences of the coding and the upstream regions of the two psbA copies were found to be identical, whereas the downstream sequences diverged from a point 20 bp 3' of the stop codons. Downstream of the gene copy designated psbAII, a dyad symmetry which allows the formation of a strong mRNA hairpin structure, and a trnH gene were found. No such elements, which are characteristic of psbA downstream regions, were found 3' of psbAI. This suggests that psbAII is the ancestral gene copy in P. contorta. Upon comparison with psbA from other plants, the pine 353-codon sequence appeared almost as distant from the angiosperm as from the liverwort counterpart. As compared to tobacco, 14 substitutions in the predicted amino acid sequence were found, most of which were located in the terminal regions of the protein.