Miseq plateform

The EasyPrep Stool Genomic DNA Kit (catalog number: DPT-BC28, TOOLS, Taipei, Taiwan) was used to extract DNA fragments from the cecum contents (0.2 g) of the mice. The samples were sequenced with on Illumina MiSeq Plateform (San Diego, CA, USA). Bioinformatics analyses of the 16S rRNA sequence data were performed to identify the bacterial species in the samples (the data were performed by Tools Microbiology Research Center, Taipei, Taiwan).

For bacterial DNA extraction, two-day protocol as described previously (http://www.openwetware.org/wiki/Gill:S._aureus_genomic_DNA_isolation), with few modifications was used. After DNA extraction, PCR amplification of 16S rRNA gene V4 region was carried out by using primers 515/806 having barcode on forward primer and sequencing using Illumina MiSeq plateform was at MR DNA (www.mrdnala.com, shallowater, TX, USA). Raw data sequences have been submitted in Sequence Read Archive and assigned an accession number PRJNA505914.

The quantity of LR-PCR DNA templates (prior and during library preparation) was assessed by spectrophotometry (Nanodrop ND-800, Thermo Scientific, Waltham, MA, USA), fluorometry (Qubit, Invitrogen, Carlsbad, NM, USA) and quantitative PCR (PicoGreen dosage with Quant-iT™ PicoGreen® dsDNA Assay Kit, Invitrogen on ABI 7900HT, Applied Biosystems). After quality controls, six libraries (one per individual) were prepared using the TruSeq DNA Sample Prep Kits v2 and TruSeq Universal Adapters (Illumina, San Diego, CA, USA). A TruSeq Universal Adapter was used for each DNA library in order to separate reads from different individuals after DNA sequencing. Library sizes were checked on BioAnalyzer chips (Agilent Technologies, Santa Clara, CA, USA). Paired-end library sequencing was carried out on the Illumina MiSeq plateform (2 × 250 bp chemistry) at the GeT-PlaGe lab (GenoToul, Toulouse, France).