16S amplicon sequencing of Southern Ocean surface seawater
Surface seawater was pumped continuously for 15 minutes through the vessels underway system (10 m depth) using an ASH 40 Allweiler peristaltic pump and Alupex hoses. Each sample corresponds to 4 L of seawater, filtered on 0.22 m cellulose ester filters (Merck Millipore model GSQP04700) with a vacuum pump (Rocker 300). DNA was extracted from the filters using the DNeasy Power Water kit (Qiagen). The V4-V5 region of the 16S rRNA gene was amplified from 1:10 diluted DNA samples, using primers 515F and 928R (Wang & Qian, 2009) and Phusion Hot Start II High-Fidelity DNA Polymerase (Thermo Scientific), with annealing temperature of 68C. PCR products were paired-end sequenced on the Illumina Miseq platform (2 x 250-bp) (DNA Sequencing Facility, University of Wisconsin Biotechnology Center, Wisconsin, USA).