title: rRNA-Seq of homo sapiens: child adenoid microbiome
design: Microbial DNA was extracted using FastDNA Spin Kit for Soil (MP Biomedicals, USA) according to the instructions of the manufacturer. FastPrep-24 Instrument (MP Biomedicals, USA) was used for sample lysis. The V3-V4 hypervariable region of the bacterial 16S rRNA gene was amplified using the 341F/805R set of primers. A unique set of oligonucleotides was added to each sample during the second amplification to ensure dual indexing (Table 1). Two negative controls were introduced during the PCR amplification, evaluated by the agarose gel electrophoresis and sequenced applying one negative control per each of two sample batches. Magnetic-bead-based purification (Macherey-Nagel, Germany) was performed for each sample after amplification. The quantity and quality of extracted DNA and amplicon libraries were determined by Qubit Fluorometer (Thermo Fisher Scientific, USA) and Agilent 2100 Bioanalyzer systems (Agilent, USA), respectively. Sequencing was done on the MiSeq System (Illumina, USA) with MiSeq Reagent Kit v2 (500-cycles) (Illumina, USA) obtaining at least 100 000 paired-end sequencing reads per sample.