Samples are diluted and run in a 30 cycle PCR that specifically amplifies the V4 region of the 16S ribosomal subunit, with the primers making use of the highly conserved regions on either side. PCR primers are barcoded by well, in a dual plate format, for a total of 192 available barcodes. Samples are quantified using the caliper, and then pooled together. Finally, the pooled sample is cleaned up and selected for size by using a combination of purification columns and a Pippen Prep SAGE protocol, and handed off for MiSeq.