Generation of Comprehensive Ecosystem-Specific Reference Databases with Species-Level Resolution by High-Throughput Full-Length 16S rRNA Gene Sequencing and Automated Taxonomy Assignment (AutoTax)
Morten Simonsen Dueholm, Kasper Skytte Andersen, Simon Jon McIlroy, Jannie Munk Kristensen, Erika Yashiro, Søren Michael Karst, Mads Albertsen, Per Halkjær Nielsen
Abstract
High-throughput 16S rRNA gene amplicon sequencing is an essential method for studying the diversity and dynamics of microbial communities. However, this method is presently hampered by the lack of high-identity reference sequences for many environmental microbes in the public 16S rRNA gene reference databases and by the absence of a systematic and comprehensive taxonomy for the uncultured majority. Here, we demonstrate how high-throughput synthetic long-read sequencing can be applied to create ecosystem-specific full-length 16S rRNA gene amplicon sequence variant (FL-ASV) resolved reference databases that include high-identity references (>98.