Amplicon, metagenome, and whole-genome sequencing
Amplicon, metagenome, and whole-genome sequencing are broadly used tools for deeply characterizing groups of microbes (who they are and what they are doing) or single microbial cells.
We offer sequence analysis services for all these technologies.
Sequencing a microbial marker gene is a fast and cost-effective tool to identify microbes from any environment such as human, animal, environmental or industrial samples. We can use it to monitor changes in groups of microbes due to changes in their surrounding environment or occurring with time.
The process begins by extracting genomic DNA from selected samples or a single microbial isolate. Next, a chosen marker gene is amplified with PCR from genomic DNA. Then the amplified genes, so-called ‘amplicons’ are DNA sequenced. For bacteria and archaea, we typically use the 16S rRNA gene. 18S rRNA gene or ITS1–ITS2 genes are a common choice for fungi.
Analysis of the sequenced 16S/18S rRNA or ITS genes begins with taxonomic annotation by mapping the gene sequences against a curated sequence database. Taxonomy annotation gives a ‘name’ for each sequence and classifies sequences into different taxonomic groups such as families and genera.
Tens of thousands of sequences per sample are easily produced from large numbers of samples to deeply characterize their microbiota.
In metagenomics or metatranscriptomics, all genomic content of a sample is sequenced. This technique is a good choice for accurately identifying microbial species and strains or their functional properties in a complex environment, having thousands of different microorganisms interacting.
The sample’s whole genomic DNA (or RNA) is randomly fragmented, and these fragments of DNA (or RNA) are sequenced. This technique is sometimes called ‘shotgun’ sequencing, referring to the pieces of sequences scattered like ‘ammo from a shotgun’ across your samples’ DNA (or RNA).
The collection of millions of sequencing reads enables identifying microbial species and strains or their functional genes by mapping reads against known genomic markers. For example, virulence genes can be important in diseases with a microbiological component.
Metagenome sequencing projects can include the assembly of sequencing reads to produce near-complete draft genomes. The assembly combines taxonomic and functional information and describes what individual organisms in the samples can do and what genes they carry.
The genome of a single microbial isolate can be sequenced, and its genes identified. Whole-genome sequencing allows thus identification of the microbial isolate in question and its properties. This can be important, e.g., in recognizing pathogenic microbes in disease outbreaks or from raw food or feed production materials.