MicroRNAs (miRNAs) are approximately 22-nt RNA segments that are involved in the regulation of protein expression primarily by binding to one or more target sites on an mRNA transcript and inhibiting translation. MicroRNAs are likely to factor into multiple developmental pathways, multiple mechanisms of gene regulation, and underlie an array of inherited disease processes and phenotypic determinants. Several computational programs exist to predict miRNA targets in mammals, fruit flies, worms, and plants. However, to date, there is no systematic collection and description of miRNA targets with experimental support. We describe a database, TarBase, which houses a manually curated collection of experimentally tested miRNA targets, in human/mouse, fruit fly, worm, and zebrafish, distinguishing between those that tested positive and those that tested negative. Each positive target site is described by the miRNA that binds it, the gene in which it occurs, the nature of the experiments that were conducted to test it, the sufficiency of the site to induce translational repression and/or cleavage, and the paper from which all these data were extracted. Additionally, the database is functionally linked to several other useful databases such as Gene Ontology (GO) and UCSC Genome Browser. TarBase reveals significantly more experimentally supported targets than even recent reviews claim, thereby providing a comprehensive data set from which to assess features of miRNA targeting that will be useful for the next generation of target prediction programs. TarBase can be accessed at http://www.diana.pcbi.upenn.edu/tarbase.