Iso-Ni 3
16S rRNA sequencing of microrganisms present in Fe(III)-reducing Ni-ISA degrading enrichment culture.
Sequencing of PCR amplicons of 16S rRNA was conducted with the Illumina MiSeq platform (Illumina, San Diego, CA, USA) targeting the V4 hyper variable regions (forward primer, 515F, 5'-GTGYCAGCMGCCGCGGTAA-3'; reverse primer, 806R, 5'-GGACTACHVGGGTWTCTAAT-3') for 2×150-bp paired-end sequencing. PCR amplification was performed using Roche FastStart High Fidelity PCR System in 50ul reactions under the following conditions: initial denaturation at 95°C for 2min, followed by 36 cycles of 95°C for 30s, 55°C for 30s, 72°C for 1 min, and a final extension step of 5min at 72°C. The PCR products were purified and normalised to 20ng each using the SequalPrep Normalization Kit . The PCR amplicons from all samples were pooled in equimolar ratios. The run was performed using a 4pM sample library spiked with 4pM PhiX to a final concentration of 10% following the method of Schloss and Kozich.
Iso-Ni 3
SUB2842629
Iso-Ni 3