A volume of 800–1,000 mL seawater was collected at the bottom of the tanks using a silicon hose fixed at the edge of the tanks to prevent contamination from the upper surface layer of oil at each sampling time (day 1, 2, 7, 14, and 28). Samples were immediately filtered through nitrocellulose filters (pore size, 0.22 μm) (Millipore, Billerica, MA, USA) and the filters were stored at −80°C until DNA extraction. DNA extraction was performed using the protocol described by Preston et al. (45 (link)) with the following modifications: the total volume of the lysate was used after filtration through a Millex-GV syringe filter (pore size, 0.2 μm; diameter, 13 mm) (Millipore) and a proportionally higher volume of diluent (555 mM sodium acetate pH 5.2 in 70% ethanol [v/v]) was added. This was then passed through a column in the DNeasy Tissue Kit (Qiagen, Hilden, Germany) and washed twice before DNA was eluted with 80 μL elution buffer.
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