Dataset: North Pond metagenome
Deployment: JRES-336

Location: North Atlantic, western flank of the mid-Atlantic Ridge 22.75589 N 46.08125 W

DNA was extracted using the method described in (Mills et al. 2012) as modified by B. K. Reese for nucleic acids in low biomass open ocean sediments. The interior of the whole-round core for sample U1382B 7H-5 was subsampled using sterile techniques and was divided into 25 splits, each weighing ~0.5 g. Initial cell lysis was achieved using five cycles of freeze (liquid nitrogen), thaw (55oC water bath) and vortex steps while stabilizing nucleic acids in a Tris–EDTA–glucose buffer. This step was followed by the addition of lysozyme to the buffer and incubation at 30oC for 10 minutes. Samples were then treated twice with buffered phenol, chloroform and isoamyl alcohol (25:24:1; pH 8.0), and sodium dodecyl sulfate to dissolve the cell membrane and solubilize both high and low molecular weight proteins. Nucleic acids within the aqueous layer above the phenol–chloroform layer were then precipitated in an ethanol and sodium acetate solution. The ethanol solution was decanted following centrifugation of samples at 4°C. Lastly, DNA pellets were resuspended in sterile water, combined into one sample. We quantified the DNA content using a Quibit fluorometer (Thermo Scientific, Waltham, MA, USA) and assessed its quality on a NanoDrop 1000 Spectrophotometer (Thermo Scientific, Waltham, MA, USA) prior to sequencing at the Marine Biological Laboratory Keck facility (Woods Hole, MA, USA). Extraction blanks were included with each sample extraction and yielded no measurable DNA.