title: Digestive gland microbiota of Mytilus galloprovincialis
design: The 16S amplicon library was prepared according to a standardized Illumina protocol (Illumina, 2013). Briefly, the V3-V4 region of bacterial 16S ribosomal RNA gene was amplified by using universal primers 341F (5- CCTACGGGNGGCWGCAG-3) and 805R (5-GACTACHVGGGTATCTAAT CC-3) (Klindworth et al., 2013) tagged with 33 bp overhang adapters. These PCR reactions (15 l) contained an initial concentration of 0.2 M of each primer, 7.5 l of Phusion High-Fidelity PCR Master Mix with HF Buffer (1X) (Thermo Scientific, USA) and 20 ng of template DNA. Triplicate PCR reactions were pooled and purified using AMPure XP beads (Beckman Coulter, USA) according to the manufacturers instructions. Secondary PCR reactions were performed in order to add Nextera XT Illumina sequencing indices and adapters. After purification, 16S amplicon libraries were pooled together at equimolar concentration and subjected to Illumina MiSeq paired-end sequencing (Illumina, USA).