16S of culture media (SE10 + TSB)
Amplicon sequencing libraries were prepared using a two-step PCR, targeting 16S rRNA gene V3-V4 regions. First PCR was performed for 30 cycles using primers Uni341F (5-CCTAYGGGRBGCASCAG-3) and Uni806R (5-GGACTACNNGGGTATCTAAT-3) originally published by Yu et al. (2005)   and modified as described by Sundberg et al. (2013). and modified as described by Sundberg et al. (2013).  First PCR amplification products were purified using HighPrep PCR clean-up (MagBio Genomics, USA) using a 0.65:1 (beads:PCR reaction) volumetric ratio. A second PCR reaction was performed to add Illumina sequencing adapters and sample-specific dual indexes (IDT Integrated DNA technologies, USA) using PCRBIO HiFi (PCR Biosystems Ltd., UK) for 15 cycles. The secondsecond PCR products were purified with HighPrep PCR Clean Up System, as described for the first PCR. Sample concentrations were normalized using the SequalPrep Normalization Plate (96) Kit (Thermofisher, USA), following manufacturers instructions. The libraries were then pooled and up-concentrated using DNA Clean and Concentrator-5 Kit (Zymo Research, USA). Library pools concentration was determined using the Quant-iT High-Sensitivity DNA Assay Kit (Life Technologies, USA) and diluted to 4 nM. The library was denatured and sequenced following manufacturers instructions, on an Illumina MiSeq platform at the Section of Microbiology - University of Copenhagen, using Reagent Kit v3 [2 x 300 cycles] (Illumina, USA).