We reconstructed the phylogeny of representatives from nine genera and three families of the Cymbellales using two nuclear and three chloroplast genes. After rooting with Anomoeoneis, Placoneis was found as sister to a clade composed of Cymbella, Cymbopleura, Encyonema, Gomphonema, and Gomphoneis. The latter group was divided into lineages with mainly heteropolar and dorsiventral valve symmetry. The data and chloroplast morphology also supported a close relationship between Geissleria decussis and Placoneis. Expectedly, the sequenced genes exhibited substantial across-site rate variation (ASRV) that prompted us to assess the stability of the inferred relationships in the face of different approaches for modelling ASRV. While the overall topology remained stable across analyses, relationships between Cymbella and Cymbopleura and within one clade of Gomphonema varied dependent on the employed model. In some cases a strongly supported relationship in one analysis was not recovered by another that differed solely in how the data were partitioned. These topological fluctuations appeared in areas of the tree with the least balanced taxon sampling, and they altered the outcomes of phylogenetic hypotheses tests of monophyly. Assessing how different models for ASRV affect tree topology and clade support values, therefore, seems important in cases of sparse or unbalanced taxon sampling or when assessing the phylogenetic affinity of previously unsampled taxa when lineage-specific biases in base composition or evolutionary rate are more difficult to detect.