Metagenomic sequencing of secondary solids from municipal wastewater
DNA extraction was carried out on the 0.5-gram solids aliquot using the FastDNA Spin Kit for Soil (MP Biomedicals, Solon, OH). Composite samples were prepared by pooling equal masses of triplicate samples. Libraries were prepared for shotgun metagenomic sequencing with the NEB Ultra II DNA Library Prep kit (New England Biolabs, Ipswich, Massachusetts) and sequenced via the Illumina NextSeq500 (Illumina, San Diego, CA) with 2x75 paired-end reads. Sequencing was performed on ten samples at the Scripps Research Institute (La Jolla, CA). Individual replicates of two samples were sequenced by both methods to assess if sequencing a pooled triplicate sample yielded an average of the three replicates.