V4 16S seq: infant Simulator of the Human Intestinal Microbial Ecosystem: distal colon
Library preparation with F515/R806 primers, Nextera XT Index v2, and gDNA. Thermocycler conditions were as follows: an initial denaturation step at 94C for 2 min; followed by 30 cycles of denaturation at 94C for 30 s, annealing at 65C for 30 s, elongation at 68C for 30s; and 10 cycles of the same parameters but with annealing at 55C, with a final extension step at 68C for 5 min. The annealing temperature comprised of a 0.3C increment touch-down starting at 65C for 30 cycles, followed by 55C for 10 cycles. PCR products purified by AMPure XP beads, normalized to 2 nM and pooled. Sequenced at the Advanced Analysis Center, University of Guelph.