Ancient DNA provides a unique means to record genetic change through time and directly observe evolutionary and ecological processes. Although mostly based on mitochondrial DNA, the increasing availability of genomic sequences is leading to unprecedented levels of resolution. Temporal studies of population genetics have revealed dynamic patterns of change in many large vertebrates, featuring localized extinctions, migrations, and population bottlenecks. The pronounced climate cycles of the Late Pleistocene have played a key role, reducing the taxonomic and genetic diversity of many taxa and shaping modern populations. Importantly, the complex series of events revealed by ancient DNA data is seldom reflected in current biogeographic patterns. DNA preserved in ancient sediments and coprolites has been used to characterize a range of paleoenvironments and reconstruct functional relationships in paleoecological systems. In the near future, genome-level surveys of ancient populations will play an increasingly important role in revealing, calibrating, and testing evolutionary processes.