Multi-Targeted Community Profiling: Leveraging the Fluidigm Access Array and Illumina MiSeq for Metagenomic Sequencing - Microorganisms

Metagenomics refers to the study of the collective set of genomes of mixed microbial communities that reside in environmental niches, soil, plant, or animal hosts.  Community profiling utilizes specific targets, such as hyper-variable regions within bacterial 16S rRNA genes, to taxonomically differentiate these communities.  The target gene is amplified by PCR using conserved priming sites and subjected to DNA sequencing. Advances in Next Generation Sequencing allows the generation of millions of amplicon sequences from indexed samples across various populations.

Sequence-based identification can be established at the genus- and species-level with the aid of sequence-classifier algorithms or alignments to specific databases that include small (16S/18S, SSU) and large subunit (23S/28S, LSU) ribosomal RNA (rRNA) sequences for bacteria, archaea and eukaryotes such as Greengenes or SILVA.

Most current amplicon assays utilize barcoded primer sets requiring the synthesis of tens to hundreds of individual primers for each region surveyed. Here, we have utilized the Fluidigm Access Array Platform for simultaneous amplification and tagging of 48 metagenomic community samples with multiple priming sets for bacterial 16S, Archeal, ITS, 18S, and specific functional genes all in a single Fluidigm run.