16S rRNA V1-V3 amplicon sequencing of vaginal microbiome
Total DNA was extracted from vaginal samples using the Qiagen DNeasy PowerSoil Kit (Qiagen, Hilden, Germany), according to the manufacturer's instructions. The V1-V3 region of the 16S rRNA gene was amplified using barcoded primers in a single-step (1-step) PCR protocol, wherein sample-specific barcodes were incorporated directly into the primers. Equimolar amounts of the resulting amplicons from each sample were pooled into a single library and sequenced on an Illumina MiSeq platform (Illumina), generating 2  300 bp paired-end reads.