The branching order of the kingdoms Animalia, Plantae, and Fungi has been a controversial issue. Using the transformed distance method and the maximum parsimony method, we investigated this problem by comparing the sequences of several kinds of macromolecules in organisms spanning all three kingdoms. The analysis was based on the large-subunit and small-subunit ribosomal RNAs, 10 isoacceptor transfer RNA families, and six highly conserved proteins. All three sets of sequences support the same phylogenetic tree: plants and animals are sibling kingdoms that have diverged more recently than the fungi. The ribosomal RNA and protein data sets are large enough so that in both cases the inferred phylogeny is statistically significant. The present report appears to be the first to provide statistically conclusive molecular evidence for the phylogeny of the three kingdoms. The determination of this phylogeny will help us to understand the evolution of various molecular, cellular, and developmental characters shared by any two of the three kingdoms. Noting that the large-subunit rRNA sequences have evolved at similar rates in the three kingdoms, we estimated the ratio of the time since the animal-plant split to the time since the fungal divergence to be 0.90.