title: 16S seq of mouse skin microbiome
design: 16S rRNA gene amplification and sequencing were carried out at the Technions Genome Center. Sequencing libraries were prepared using the 16S rRNA Metagenomic Sequencing Library Preparation protocol by Illumina with minor adjustments. Sample input was 0.625 ng of genomic DNA and the first PCR amplification consisted of 30 cycles. V3 and V4 hypervariable regions of bacterial 16S rRNA were the amplification targets. Primers used to target the V3 and V4 regions of the 16S rRNA gene were 5'TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGCCTACGGGNGGCWGCAG (Forward) and 5'GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGACTACHVGGGTATCTAATCC (Reverse). The respective Illumina overhang adapter sequences were included in the primerdesign. All 96 libraries were sequenced on an Illumina MiSeq instrument with 250 paired-ends reads. Sequencing data was input into the 16S rRNA gene Metagenomics app on the Illumina BaseSpace sequence hub. Classification was performed using the Illumina 16S rRNA gene Metagenomics workflow, which includes demultiplexing of indexed reads, FASTQ files generation, and read classification. OTU clustering and classification were performed at genus and species level. An Illumina-curated version of the Greengenes database was used as thC taxonomy database for the metagenomics workflow. The algorithm used is a high-performance implementation of the Ribosomal Database Project Classifier described in Wang Q. et al. [43].