Sampling, extraction, and high-throughput sequencing methods for environmental microbial and viral communities

Abstract

The emergence of high-throughput sequencing technologies has deepened our understanding of complex microbial communities and greatly facilitated the study of as-yet uncultured microbes and viruses. Studies of complex microbial communities require high-quality data to generate valid results. Here, we detail current methods of microbial and viral community sample acquisition, DNA extraction, sample preparation, and sequencing on Illumina high-throughput platforms. While using appropriate analytical tools is important, it must not overshadow the need for establishing a proper experimental design and obtaining sufficient numbers of samples for statistical purposes. Researchers must also take care to sample biologically relevant sites and control for potential confounding factors (e.g., contamination).