title: FS853 archaeal v6 survey
design: "Construction protocol: The v6 hypervariable region of the archaeal SSU rRNA gene was amplified using degenerate primers. The primers were 958F (AATTGGANTCAACGCCGG) and 1048R (CGRCRGCCATGYACCWC). The forward primers included 5 nt multiplexing barcodes and the reverse a 6 nt index. We generated PCR amplicons in triplicate 33 uL reaction volumes with an amplification cocktail containing 10.0 U Platinum Taq Hi-Fidelity Polymerase (Invitrogen, Carlsbad, CA), 1X Hi-Fidelity buffer, 200 uM dNTP PurePeak DNA polymerase mix (ThermoFisher), 2 mM MgSO4 and 0.3 uM of each primer. We added approximately 10-25 ng template DNA to each PCR and ran a no-template control for each primer pair. Amplification conditions were: initial 94C, 3 minute denaturation step; 25 cycles of 94C for 30s, 60C for 45s, and 72C for 60s; final 2 minute extension at 72C. The triplicate PCR reactions were pooled after amplification and purified using Qiagen MinElute plates followed by clean up, PicoGreen quantitation and Sage PippinPrep size selection."