salamander environment metageome
We processed skin samples using the protocol outlined in Hernndez-Gmez et al. (2017). To isolate microbial DNA from skin swabs, we used the DNeasy PowerSoil Kit (Qiagen N.V., Hilden, Germany). To control for contamination, we included negative extraction controls (i.e., unused swabs) and researcher glove swabs. We amplified the bacterial 16S rRNA gene V4 region using primer pair F515/R806 attached to barcode/sequencer adaptor connector sequences. We ran each sample in triplicate, with each 15-L reaction containing 5 L of template DNA, 7.5 L of 2X MyTaq Master Mix (Bioline, Tauton, MA, USA), 1.0 L of 1 nM forward and reverse primers, and 1.5 L of sterile water. PCR conditions consisted of 94C for 3 min, 30 cycles of 94C for 45s, 50C for 60s, and 72C for 90s, followed by 72C for 10 minutes. We pooled triplicate PCR products and cleaned them using the Qiagen UltraClean PCR Clean-up kit.