As the cost of sequencing declines, more and more laboratories are extending their research into metagenomics. The Metagenomics RAST Server (MG-RAST) is one of the leading tools for metagenomic analysis, and in January’s issue of Cold Spring Harbor Protocols, Folker Meyer and colleagues from the Argonne National Laboratory present a detailed set of instructions for using this web-based open source system, Using the Metagenomics RAST Server (MG-RAST) for Analyzing Shotgun Metagenomes. Shotgun metagenomics creates millions of fragments of short DNA reads, which are meaningless unless analyzed appropriately. MG-RAST offers a unique suite of tools for analyzing these data sets. After dereplication and quality control, fragments are mapped against a comprehensive nonredundant database (NR). Phylogenetic and metabolic reconstructions are computed from the set of hits against the NR. The resulting data are made available for browsing, download, and most importantly, comparison against a comprehensive collection of public metagenomes.