Assembly

Samples were collected from three timepoints during the incubation of the digestate: before N2O was added, and after 115 and 325 hours with N2O inclusion. Additionally, samples from a pre-treated digestate (heat-treated at 55 °C for 2 hours before incubation with N2O) was included to increase the the differential coverage for improved binning. DNA was extracted (PowerLyzer® PowerSoil® DNA Isolation kit from QIAGEN, following the kit protocol) and and sequenced (TruSeq PCR-free library preparation, HiSeq4000 150PE sequencing).The generated raw reads were trimmed with Trimmomatic (v0.36) before co-assembly with metaSPAdes (v3.10.1). Short contigs (<500 bp) are excluded before the generated contigs were subjected to downstream analysis (i.e. binning and functional annotation).