16S rRNA gene amplicon sequencing of biofilm DNA
PCR primers targeting Bacteria and Archaea were used for amplification of the V4 region of 16S rRNA genes with forward primer 515F (Caporaso) and barcoded reverse primer 806R (Caporaso) according to Earth Microbiome Project Protocols. The forward primer was linked to Illumina adapter as follows: 5' Illumina adapter - Forward primer pad - 2bp linker - Forward primer, 515FF (515F: AATGATACGGCGACCACCGAGATCTACAC TATGGTAATT GT GTGCCAGCMGCCGCGGTAA). The reverse primer additionally included a unique 12 bp barcode as follows: Reverse complement of 3' Illumina adapter - 12 bp Golay barcode - Reverse primer pad - Reverse primer linker - Reverse primer, 806R (806R: CAAGCAGAAGACGGCATACGAGAT XXXXXXXXXXXX AGTCAGTCAG CC GGACTACHVGGGTWTCTAAT)