16S rRNA gene amplicon sequencing of water column DNA
PCR primers targeting Bacteria and Archaea were used for amplification of the V4 region of 16S rRNA genes with  forward primer 515F (Parada et al. 2015, doi: 10.3354/ame01753) and reverse primer 806R (Apprill et al. 2015, doi: 10.3354/ame01753) according to Earth Microbiome Project Protocols except that PCR primers were dual-barcoded following Kozich et al. 2013 (doi: 10.1128/AEM.01043-13). The forward primer was linked to an Illumina adapter as follows: 5' Illumina adapter - 8 bp barcode - Forward primer pad - 2bp linker - Forward primer 515F (515F: AATGATACGGCGACCACCGAGATCTACAC-NNNNNNNN-TATGGTAATT-GT-GTGYCAGCMGCCGCGGTAA). The reverse primer was linked to an Illumina adapter as follows: Reverse complement of 3' Illumina adapter - 8 bp barcode - Reverse primer pad - Reverse primer linker - Reverse primer 806R (806R: CAAGCAGAAGACGGCATACGAGAT-NNNNNNNN-AGTCAGTCAG-CC-GGACTACNVGGGTWTCTAAT)