Genome Sequencing of Evolved Nitrite Strains

We sequenced the genomes of evolved isolates using methodology described elsewhere [20 (link)]. Briefly, we streaked each evolved nitrite (NO₂⁻) cross-feeding co-culture onto LB agar plates containing 10 μg ml^− 1 of gentamicin and 0.1 mM of IPTG and picked a single colony of the nitrite producing and reducing strain (each colony expressed a different fluorescent protein) from each co-culture for genome sequencing. We grew the single clones in LB medium overnight and extracted the DNA with a Wizard Gemoic DNA purification kit (Promega, Madison, WI). We then sent the extracted DNA to the ETH Quantitative Genomics Facility (Basel, Switzerland) for sequencing. The genomes were sequenced with an Illumina HiSeq 200 sequencer (Illumina, San Diego, CA) with 100 cycles of paired-end sequencing. Primary data analysis, de-multiplexing and quality control analysis of the sequencing data were performed using FastQC (Illumina, San Diego, CA). We reported the complete set of parameters used for quality control elsewhere [17 (link)].