16S rRNA sequencing of human gut microbiome: Ctrl-pH6.5 3 d4.7 Illumina
Microbial DNA was extracted from human fecal-derived bioreactor samples. The V4 region (~291 bp) of the 16S rRNA gene was amplified using the barcoded 515F/806R primer set following the Earth Microbiome Project (EMP) protocol and Illumina 16S Metagenomic Library Prep Guide. A two-step PCR workflow was used, including barcoding and library indexing. Amplicons were pooled, purified with KAPA Pure beads, normalized to 1 nM, and sequenced on an Illumina NextSeq 2000 platform using 2  150 bp paired-end reads. Library QC was performed using Qubit, and read quality was assessed using FastQC and MultiQC.