This dissertation focuses on increasing the understanding of the evolution processes that have contributed to the diversification of freshwater crayfish. Chapter one estimates the divergence time of the three crayfish families and tests the hypothesis that diversification is tied to the break-up of Pangaea, Gondwanna, and Laurasia. I find that the families of crayfish diverged prior to or in association with the break-up of the three super continents. Chapter two addresses the evolutionary history of the genus Cambarus, using molecular data to test hypotheses of relationships based on chela and carapace morphology. The results provide evidence that the morphology used to determine Cambarus relationships do not reflect evolutionary history and that convergent evolution of morphological traits is common in crayfish. Chapter three addresses evolution at the population level and tests for differences in the genetic population structure of two crayfish with different physiological needs. I find that physiological requirements of these crayfish have influenced their population genetic structure. The last chapter addresses a molecular based hypothesis that rates of mitochondrial evolution are reduced in cave crayfish that have increased longevity, reduced metabolism, and restricted diets compared to surface crayfish. I find that cave crayfish rates of mitochondrial evolution do not significantly differ from surface crayfish. Therefore, increased longevity, reduced metabolism, and restricted diets do not slow the rate of mitochondrial evolution as predicted in this group of cave crayfish.