Recent phylogenetic investigations of Cupressoideae have found evidence to suggest that Cupressus is not monophyletic. This study tested the division of Cupressus into an Old World clade and a New World clade with complete sampling of the 28 extant species. Data from anatomy,
biochemistry, micromorphology, reproductive development, reproductive morphology, and vegetative morphology were combined with molecular sequence data (matK, NEEDLY intron 2, nrITS, rbcL, and trnL) to produce the most complete hypothesis of evolutionary relationships within
Cupressoideae to date. Callitropsis, Cupressus, and Juniperus formed a well–supported monophyletic group (100%). Within this clade, the only demonstrably monophyletic genus was Juniperus (100%). Monophyly of the 12 Old World species of Cupressus was
well supported (100%). Old World species of Cupressus were sister to Juniperus (99%). Callitropsis and the 16 New World species of Cupressus were resolved as the sister group to the Old World Cupressus plus Juniperus clade (100%), rendering Cupressus
polyphyletic. The relationship between Callitropsis and the New World species of Cupressus was not resolved. Based on the results of the combined analysis, generic circumscriptions were modified: Cupressus was restricted exclusively to Old World species and Callitropsis
was expanded to include the New World species previously classified as Cupressus (seventeen new combinations in Callitropsis were made).