Microbiome_4448N
16S_metagenomics
Dual-indexed universal primers were utilized for the two-steps PCR amplification of the V3/V4 hypervariable regions of the 16s rRNA gene. The first-step PCR amplicons were then used for the second-step PCR for further amplification and inclusion of indexes (Nextera XT Index Kit v2 Set A).  The resulting PCR products were quantified by Qubit™ 1X dsDNA HS Assay Kit, pooled in equimolar amounts and then purified with AMpure-XP magnetic beads. The final library containing all the pooled samples was sequenced with version 3 of MiSeq Reagent Kit, 2x300 bp output, on a MiSeq system.