title: 16S v3v4 of Equus caballus: adult female feces
design: Using 2 mL bead beating tubes (0.7 mm Dry Garnet) and a Vortexed-Genie 2 fitted with Qiagens Vortex Adapter (cat. No. 13000-V1-24), we homogenized 0.20-gram sub-samples of horse fecal material. We extracted DNA from homogenized fecal samples using QIAgens QIAamp PowerFecal DNA Kits, following manufacturer recommendations outlined in the Qiagen PowerFecal DNA handbook. Notably, we used a single tube extraction protocol (rather than a 96-well format) and randomized the order in which samples were extracted. In the final step, we eluted DNA from the spin columns using 100 l of ddH2O pre-warmed to 60C. Prior to sequencing, we quantified the DNA in eluted extracts using a Qubit dsDNA BR Assay Kit and standardized DNA concentration to 20 ng/uL prior to PCR amplification. We PCR amplified the v3v4 region of the 16S rRNA gene using the 341f forward and 805r reverse universal primers. PCR products were sequenced on an Illumina MiSeq platform  (v3 chemistry: 2 x 300 base-pair read pairs) at the University of Calgary Centre for Health Genomics and Informatics.