METAGENOMIC ANALYSIS

Metagenomics is the study of microbial communities sampled directly from their natural environment, without prior culturing.

By enabling analysis of populations – including many unculturable and often unknown microbes – metagenomics is revolutionizing the field of microbiology. Advances in this field could greatly benefit research in various disciplines, including those in ecology, environmental sciences, and biomedicine. Currently, metagenomics projects are facilitated by the rapid development of NGS techniques, which provide lowered experimental costs inherent in conventional sequencing methods.

Mass sequencing of uncultured, unpurified microbial or viral populations, followed by bioinformatics analyses permits the identification and characterization of unknown pathogens necessary for clinical, medicinal and ecosystem applications. Metagenomic approaches in medicine are poised to deliver information of high diagnostic value for infectious diseases and many chronic health conditions.

Metagenomic analysis falls into two broad categories. Large-scale shotgun metagenomics typically involve extensive sequencing of the entire meta-genome in the sample, followed by de novo partial assembly of individual genomes using appropriate algorithms. Targeted metagenomics, as the name suggests, typically aim at smaller-scale goals, such as the 16S rRNA-based surveys for microbe identification, or acquiring sequence reads with specific protein functions, such as antibiotic resistance genes. At ACGT we offer both types of analysis.

How targeted metagenomic analysis works

Total DNA is extracted from the sample, followed by various selected PCR reactions. The short PCR products (550 bp maximum size) are purified and are converted to Illumina® libraries by a second stage PCR with primers containing Illumina indices and adapters. Dual indexing is typically used to maximize sample processing efficiency (See Important Considerations and Guidelines for Sample Submission). The amplicon libraries are analyzed using long paired-end reads to fully sequence each PCR product in the cluster. The collapsed reads are sorted, counted, and non-identical sequences are BLASTed against selected databases for sequence identification.