WATER5
The V1-V9 region of the 16S rRNA gene (27-1492 nt) from bacteria of water samples was sequenced using the Oxoford Nanopore MinION device, and the sequencing kit 16S Barcoding 1-24 (SQK-16S024). An input of 50 ng of DNA were used for the PCR amplification, using the LongAmp Hot Start Taq 2 Master Mix (NEB, M0533S) and the primers provided in the kit (F: 5 - ATCGCCTACCGTGAC - barcode - AGAGTTTGATCMTGGCTCAG - 3 and R: 5 - ATCGCCTACCGTGAC - barcode - CGGTTACCTTGTTACGACTT - 3). Cycling conditions were 95 C for 1 min, followed by 30 cycles of 95C for 20 s, 52C for 30 s, and 65C for 2 min with a final extension step of 65 C for 5 min (Toxqui-Rodriguez et al., 2023). After a clean-up step using AMPure XP Beads (Beckman Coulter), amplicons were quantified using PicoGreen dye (Thermo Fisher Scientific). Samples were pooled in 100 fmol pools and loaded in MinION devices using R9.4.1 Flow Cells (FLO-MIN106D).