A dated molecular phylogeny is proposed for the Tanypodinae, a diverse subfamily of Chironomidae (Diptera). We used molecular data from fragments of one ribosomal gene (28S), one nuclear protein-coding gene (CAD), and one mitochondrial protein-coding gene (COI), analysed using mixed model Bayesian and maximum likelihood inference methods. All proposed tribes were sampled, namely, Anatopyniini, Clinotanypodini, Coelopyniini, Fittkauimyiini, Macropelopiini, Natarsiini, Pentaneurini, Procladiini and Tanypodini. A multilocus dataset of 1938 characters was compiled from 123 individuals including outgroups. Monophyly was supported for all tribes although some relationships were not robust. Relationships between tribes and some genus groups are highly congruent with a morphology-based estimate. Relationships within tribe Pentaneurini mostly find weak support, yet previously hypothesised groupings and monophyly or lack thereof in well-sampled genera are revealed. The tempo of diversification of the family was deduced by divergence time analysis (BEAST). Origination of a subfamily stem group in the late Jurassic to early Cretaceous was inferred, with all tribes and many genera of Pentaneurini originating and diversifying in the Cretaceous. Some nodes are biogeographically informative. Gene sections supported the backbone, but more extensive sampling is needed to estimate shallower phylogenies and to better understand the tempo and diversification of Tanypodinae.