Whole genome shotgun sequecning of Human Vagina
Metagenomic libraries were prepared using the KAPA HyperPlus Kit (Kapa Biosystems) with KAPA Single-Indexed Adapter Kit Set B. A fixed volume (35 µL) of genomic DNA was used as input, and libraries were prepared following the manufacturer's protocol with modifications based on their amount of input DNA. For samples with 0.5 or 0.2 ng input DNA, the fragmentation enzyme was diluted 1:2 or 1:5 with water. All samples were fragmented at 37oC for 5 min. The post-ligation cleanup was performed with 0.8x Ampure XP beads (Beckman Coulter, Indianapolis IN) and 20 µL of sample was used in library amplification. Amplification library cycles varied by input DNA. Post-amplification cleanup was performed with 1x Ampure XP beads; libraries with remaining adapter dimer peaks were cleaned a second time. The final elution was in 25 µL of nuclease-free water. Libraries were run on a TapeStation instrument with a D1000 tape (Agilent) to assess quality and concentration. Libraries were sequenced (8 libraries/lane) on an Illumina HiSeq 4000 instrument using the 150 bp paired-end protocol. Host reads were removed from all metagenomic sequencing data using BMTagger and the GRCh38 reference genome, and reads were quality filtered using trimmomatic (v0.38, sliding window size 4bp, Q15, minimum read length:75bp).