The collection of papers in this issue of Mycologia documents considerable improvements in taxon sampling and phylogenetic resolution regarding the Fungal Tree of Life. The new data will stimulate new attempts to date divergences and correlate events in fungal evolution with those of other organisms. Here, we review the history of dating fungal divergences by nucleic acid variation and then use a dataset of 50 genes for 25 selected fungi, plants and animals to investigate divergence times in kingdom Fungi. In particular, we test the choice of fossil calibration points on dating divergences in fungi. At the scale of our analysis, substitution rates varied without showing significant within-lineage correlation, so we used the Langley-Fitch method in the R8S package of computer programs to estimate node ages. Different calibration points had a dramatic effect on estimated divergence dates. The estimate for the age of the Ascomycota/Basidiomycota split was 1808000000 y ago when calibrated assuming that mammals and birds diverged 300000000 y ago, 1489000000 y ago when calibrated assuming that the 400000000 y old fungal fossil Paleopyrenomycites devonicus represents Sordariomycetes and approximately 400000000 y ago when calibrated assuming 206000000 y ago for the plant eudicot/monocot divergence. An advantage of a date of approximately 400000000 y ago for the Ascomycota/Basidiomycota divergence is that the radiation of fungi associated with land plants would not greatly precede the earliest land plant fossils. Acceptance of approximately 400000000 y ago for the Ascomycota/Basidiomycota split would require that P. devonicus be considered a deeply branching Ascomycota. To improve on current estimates of divergence times, mycologists will require calibration points from within groups of fungi that share similar substitution rates. The most useful calibration is likely to depend on the discovery and description of continuous records of fossil fungi, or their spores, that show recognizable shifts in morphology.