Uncovering the Diversity of Pathogenic Free-Living Amoebae in Freshwater Environments of Ghana: A Combined Culture Enrichment and Metabarcoding Approach
Environmental DNA (eDNA) samples were collected from freshwater stream water to assess eukaryotic community diversity. The 18S rDNA gene was targeted using PCR amplification with primers specific to the V3V4 region, including CCGCGGTAATTCCAGCTC (forward) and CCCGTGTTGAGTCAAATTAAGC (reverse), based on Hadziavdic et al. (2014). High-fidelity Phusion polymerase was used, and PCRs were performed across a gradient of annealing temperatures (5060C) to maximize eukaryotic taxon recovery. Amplicons were purified, quantified using Qubit dsDNA BR assay, and pooled. Library preparation was performed using the Oxford Nanopore Rapid Barcoding Kit (SQK-RBK114.24), and sequencing was carried out on a MinION device using an R10.4.1 flow cell. This study aims to characterize microbial eukaryotic diversity from freshwater ecosystems using long-read metabarcoding.
SUB15428582
Metagnomics_Ghana_Suruwi_3