The level of chloroplast DNA (cpDNA) diversity was estimated among individuals, populations, and geographic regions of Douglas-fir (Pseudotsugamenziesii(Mirb.) Franco) in British Columbia by analyzing restriction fragment length polymorphisms (RFLP) from 18 populations covering the three geographic regions (coast, transition zone, and interior). Four sets of pooled probes, consisting of four to eight cpDNA clones from lodgepole pine (PinuscontortaDougl. ex Loud.), were used as hybridization probes for RFLP analysis. Thirty cpDNA restriction fragments out of a total of 175 observed fragments were detected to be polymorphic in all four sets of hybridization probes. Sixteen haplotypes were characterized among all of the Douglas-fir populations. The transition region showed the highest level of total genetic diversity (0.853) and genetic diversity within populations (0.762) of the three regions. The highest degree of population differentiation was found in the interior region (0.193). A higher proportion of the genetic diversity was allocated within populations than among populations in all three regions. Only 11–19% (mean 14.6%) of the total genetic diversity within geographic regions is due to interpopulation genetic diversities. The distribution of cpDNA diversity in sampled B.C. Douglas-fir is concordant with a typical pattern for long-lived woody species as observed in allozyme studies.