16S rRNA V3-V4 amplicon sequencing of normal control mouse gut microbiota - replicate 4 (NC_4)
16S rRNA gene V3-V4 hypervariable region amplicon sequencing of mouse gut microbiota. Genomic DNA (30 ng) was amplified using fusion primers targeting the 16S V3-V4 region. PCR products were purified using Agencourt AMPure XP magnetic beads and quality-checked using Agilent 2100 Bioanalyzer. Libraries were sequenced on BGISEQ-500 platform (paired-end, 300 bp reads). Raw reads were filtered using cutadapt v2.6 and quality-trimmed with a 30 bp sliding window (Q>20); reads shorter than 75 percent of original length discarded. Tags assembled using FLASH v1.2.11 (min overlap 15 bp, mismatch rate 0.1). OTUs clustered at 97 percent similarity using USEARCH v11.0.667; chimeras removed using UCHIME v4.2.40 against gold database. Taxonomy assigned using RDP classifier v1.9.1 against RDP Release19 database (confidence threshold 0.6). BGI Project Code: F24A040009445_MUSqcjhM.