Technical Abstract: Comparative analysis of multiple genomes in a phylogenetic framework dramatically improves the precision and sensitivity of inferences in evolutionary genomics. The genomes of 12 Drosophila species, nine of which are presented here for the first time (sechellia, yakuba, erecta, ananassae, persimilis, willistoni, mojavensis, virilis and grimshawi), illustrate how rates and patterns of sequence divergence can be used to gain insight into evolutionary processes on a genomic scale. These genome sequences augment the remarkable genetic tools that have made Drosophila melanogaster a pre-eminent model for animal genetics, and will further catalyze fundamental research on mechanisms of development, cell biology, genetics, disease, neurobiology, behaviour, physiology, and evolution. Comparative genomic data allow many internal checks on data quality, producing higher confidence in robust results than single genome analyses can provide. Despite the remarkable similarities among these Drosophila species, we identified many lineage-specific changes in protein-coding genes, noncoding RNA genes, and cis-regulatory regions that deviate from strict neutrality and may prove to underlie differences in ecology and behaviour of these diverse species.