A restriction-site analysis of chloroplast DNA from 44 species, representing 19 genera and all six subfamilies of the Crassulaceae was conducted using 12 restriction endonucleases. A total of 969 variable sites was detected, 608 of which were phylogenetically informative and used in parsimony analysis. Estimated values of nucleotide sequence divergence were used to construct a distance tree by the neighbor-joining method. Maximum sequence divergence in the family was ~7%. Different tree inference methods yielded only moderately different topologies. The amount of support for the monophyletic groups obtained in the Wagner parsimony analysis was evaluated by bootstrap and decay analysis. There is very strong support for a basal division of the family, which separates the monophyletic subfamily Crassuloideae from all other taxa. Four of the six traditionally recognized subfamilies are indicated to be polyphyletic. These include the Cotyledonoideae, Sempervivoideae, Sedoideae, and Echeverioideae. The Kalanchoideae and the genera Cotyledon and Adromischus exhibit low levels of cpDNA sequence divergence relative to one another, suggesting a relatively recent radiation. The genera Sedum and Rosularia are indicated to be polyphyletic. Sedum comprises sister taxa of most of the other genera of the family.