In this review, we highlight the importance of reference sets in human microbiome research, limitations of existing resources, technical challenges to employing reference sets, examples of prototypical reference usages, and contributions of the American Gut Project to addressing some of these issues. Discussion will focus on the 16S ribosomal RNA (16S rRNA) gene, which is a popular locus for use in microbiome studies over a wide range of environment types [19–23] and is the core locus assayed in the American Gut Project. Construction of references based on other loci is important for studying microbial eukaryotes, viruses, and interactions between these organisms, but high-throughput study of these other components of the community is not yet cost-effective.
