Dataset: Adelie Basin 16S Amplicons
Deployment: JRES-318

Samples were collected at 66.413S, 140.425E from depths 0.05 - 97 meters below sea floor.

Samples for molecular biological analysis were collected shipboard from 10 cm sections of the intact core using sterile cut-end 5 mL syringes. Samples were immediately frozen at –80 degrees C and maintained at this temperature during transport and storage to the home laboratory. Genomic DNA was extracted from sediment samples using a bead beating/phenol chloroform protocol. Amplicons of the 16S rRNA gene were prepared for 454 pyrosequencing using a PCR-touchdown annealing temperature strategy from Don et al. (1991) with procedure modifications and modified universal primers 515f and 927r of Osburn et al. (2011). Forward primers included the 454 Life Science A adaptors and a sample specific 8 nt barcode. The reverse primer included the 454 Life Science B adaptors. Amplicon concentrations for each sample were quantified using a 2100 Bioanalyzer (Agilent Technologies, Colorado Springs, CO, USA), pooled (22 ng DNA/sample) and concentrated using a Savant DNA 12 Speed Vac Concentrator (Thermo Scientific, Waltham, MA, USA). The pooled DNA was gel purified using the Montage DNA Gel Extraction Kit (Millipore, Bellerica, MA, USA), and sequenced on a Roche 454 FLX titanium platform at Engencore, University of South Carolina (now Selah Genomics).