We have constructed extensive 18S-28S rDNA dinoflagellate phylogenies (>200 sequences for each marker) using Maximum Likelihood and Bayesian Inference to study the evolutionary relationships among marine and freshwater species (43 new sequences). Our results indicated that (a) marine and freshwater species are usually not closely related, (b) several freshwater species cluster into monophyletic groups, (c) most marine-freshwater transitions do not seem to have occurred recently and, (d) only a small fraction of the marine lineages seem to have colonized fresh waters. Thus, it becomes apparent that the marine-freshwater boundary has acted as a barrier during the evolutionary diversification of dinoflagellates. Our results also shed light on the phylogenetic positions of several freshwater dinoflagellates which, to date, were uncertain.