Whole Genome Sequencing of Feces collected from Healthy male Sigmodon hispidus
A subset of fecal samples from 20 total male cotton rats (10 from each species), taken at days 34 and 111 within the first cohort of cotton rats, underwent whole-metagenomic shotgun sequencing. From the same stool samples, genomic DNA was extracted using the Qiagen DNeasy PowerSoil Kit (Cat No./ID: 12888100) by following the manufacturers protocol (skipping the optional 4C incubations). In addition, a negative sample (which did not contain any template but was otherwise processed the same as the rest of the samples) and a positive control (ZymoBIOMICS Microbial Community Standard) were processed in parallel with samples and sequenced. Samples were normalized to 75ng/ L in 1X TE prior to library construction. Metagenomic libraries were prepared using the NEBNext Ultra II FS DNA Library Prep Kit for Illumina following the manufacturers protocol for inputs 100ng. Samples were fragmented at 37C for 12min to yield a fragment size of 200450bp. NEBNext Multiplex Adaptors were diluted 10-fold. NEBNext Multiplex Oligos for Illumina (Set 1, NEB #E7335) were used for PCR enrichment of adaptor-ligated DNA, and 5cycles of PCR were run. Library quality was assessed on an Agilent 2100 Bioanalyzer System using the Agilent High Sensitivity DNA Kit (50674626). Samples were sequenced via the NovaSeq 6000 2150 platform for Illumina at the Vanderbilt Technologies for Advanced Genomics (VANTAGE) core, aiming for 40 million reads per sample.