DNA was extracted from triplicate enrichments grown in 50 ml of media with 5 g of Fe0 as the sole electron donor. Extractions were only carried out on the entire corrosive community (untreated with inhibitors) at the 1-month mark of the 4th transfer (for shotgun metagenome sequencing) and 11th transfer (for amplicon sequencing), when both acetogens and methanogens are sufficiently active according to physiology data.
Before metagenome sequencing, genomic DNA was isolated from the pellets of triplicate enrichments (fourth transfer) using commercially available kits, as previously described (Palacios et al., 2019 (link)).
Before amplicon sequencing, genomic DNA was isolated from the cell filtrates of triplicate enrichments (11th transfer) using a FastDNA Spin kit for Soil (MP Biomedicals, United States) with the following modifications: to the Lysing Matrix E tube, we added 500 μl of sample, 480 μl of sodium phosphate buffer, and 120 μl of MT buffer. Bead beating was performed at 6 m/s for 4 × 40 s (Albertsen et al., 2015 (link)).
The integrity of genomic DNA was verified on an agarose gel and quantified on a mySPEC spectrophotometer (VWR®/Germany) or Qubit dsDNA HS/BR Assay kit (Thermo Fisher Scientific, United States).
Before metagenome sequencing, genomic DNA was isolated from the pellets of triplicate enrichments (fourth transfer) using commercially available kits, as previously described (Palacios et al., 2019 (link)).
Before amplicon sequencing, genomic DNA was isolated from the cell filtrates of triplicate enrichments (11th transfer) using a FastDNA Spin kit for Soil (MP Biomedicals, United States) with the following modifications: to the Lysing Matrix E tube, we added 500 μl of sample, 480 μl of sodium phosphate buffer, and 120 μl of MT buffer. Bead beating was performed at 6 m/s for 4 × 40 s (Albertsen et al., 2015 (link)).
The integrity of genomic DNA was verified on an agarose gel and quantified on a mySPEC spectrophotometer (VWR®/Germany) or Qubit dsDNA HS/BR Assay kit (Thermo Fisher Scientific, United States).
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