title: Shotgun metagenomic sequencing of Dactylorhiza traunsteineri rhizosphere microbiome - Soil
design: Isolation of the metagenomic DNA was performed from the rhizosphere soil and the plantlet root wash lyophilized samples in a sterile environment. Shotgun metagenomic libraries were prepared using 1000 ng of genomic DNA, which was fragmented using the Bioruptor sonication device. Fragments were cleaned up using the GeneJet PCR Purification Kit and size-selected for a 300 bp using Agencourt AMPure XP beads. Sequencing libraries were prepared following the standard NEBNext Ultra II DNA Library Prep with Sample Purification Beads procedure. The final library concentrations were determined using the Qubit dsDNA HS Assay Kit and the average size of the library was determined using the Agilent 5200 Fragment Analyzer System. The libraries were pooled and diluted to 4 nM before being sequenced for 600 cycles pair-end on the Illumina MiSeq system using the MiSeq Reagent Kit v3.