river biofilm 16S amplicon sequencing
The V3-V4 variable regions of the 16S rRNA gene were amplified in a one-step PCR using the primer pair 341F-806R (dual-barcoding approach). After verification of the presence of PCR-products by gel electrophoresis, normalization (Sequal Prep Normalization Plate Kit; Thermo Fisher Scientific, Waltham, USA) and equimolar pooling was performed. Sequencing was conducted on the MiSeq platform (MiSeq; Illumina, San Diego, USA) with v3 chemistry (2x300 bp). The settings for demultiplexing were 0 mismatches in the barcode sequences.