16S rRNA sequencing ofN2O reducing chemostat enrichment: day 34
DNA was extracted from a pellet retrieved from 2 mL of the enrichment culture at each sampling occasion using the PowerLyzer PowerSoil DNA Isolation Kit (MoBio Laboratories). The 16S rRNA genes were amplified in duplicate using 25+8 cycles and the universal primers pro341F and pro805R (Takahashi et al., 2014). The PCR reactions contained 10 ng template DNA, Phusion High-Fidelity PCR Master Mix (Thermo Scientific), 0.75  g/ l BSA and 0.25  M of each primer, which were equipped with Nextera adapter sequences in the second PCR. Amplicons were pooled and purified with the AMPure Beads Purification kit (Agilent Technologies) in between and after the final amplification step. Sequencing was performed by Microsynth (Microsynth AG, Switzerland) on the MiSeq platform (Illumina) using 2x300 bp paired-end chemistry.
N2Oreducing_chemostat_day34