Partial sequences of nuLSU rDNA were obtained to investigate the phylogenetic relationships of Pyronemataceae, the largest and least studied family of Pezizales. The dataset includes sequences for 162 species from 51 genera of Pyronemataceae, and 39 species from an additional 13 families of Pezizales. Parsimony, ML, and Bayesian analyses suggest that Pyronemataceae is not monophyletic as it is currently circumscribed. Ascodesmidaceae is nested within Pyronemataceae, and several pyronemataceous taxa are resolved outside the family. Glaziellaceae forms the sister group to Pyronemataceae in ML analyses, but this relationship, as well as those of Pyronemataceae to the other members of the lineage, are not resolved with support. Fourteen clades of pyronemataceous taxa are well supported and/or present in all recovered trees. Several pyronemataceous genera are suggested to be non-monophyletic, including Anthracobia, Cheilymenia, Geopyxis, Humaria, Lasiobolidium, Neottiella, Octospora, Pulvinula, Stephensia, Tricharina, and Trichophaea. Cleistothecial and truffle or truffle-like ascomata forms appear to have evolved independently multiple times within Pyronemataceae. Results of these analyses do not support previous classifications of Pyronemataceae, and suggest that morphological characters traditionally used to segregate the family into subfamilial groups are not phylogenetically informative above the genus level.