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Full-Length 16s rRNA / ITS Metagenomic Sequencing Service

What is 16S rRNA sequencing?

16S rRNA gene sequencing is commonly used for identification, classification and quantitation of microbes within complex biological mixtures such as environmental samples (example marine water) and gut samples (example human gut microbiome).Conveniently, the 16S rRNA  ene consists of both conserved and variable regions.

Why is the 16S rRNA sequence used for identification?

Because of the complexity of DNA–DNA hybridization, 16S rRNA gene sequencing is used as a tool to identify bacteria at the species level and assist with differentiating between closely related bacterial species. Many clinical laboratories rely on this method to identify unknown pathogenic strains.

Why choose full length 16S rRNA sequencing over partial length 16S rRNA sequencing?

The complete length of 16S rRNA gene is around 1500-1600 bp allowing for more information to be utilized for species identification. Meanwhile, partial length uses a short fragment of the gene, usually around 300-500 bp, meaning not all information can be obtained. Limited by the information availability, partial-length method can only be used to identify the genus name of bacteria. In contrast, the full-length method can be used to identify the species name. Genetika Science Indonesia is offering full length amplicon sequencing for metagenomics using GridION from Oxford Nanopore Technologies (ONT).

 

Platform : ONT GridION system
Sample Type:
1. Raw Sample
2. Genomic DNA

  • For genomic DNA samples:
  • Amount: 100 ng (Nanodrop)
  • Volume : 30 μL
  • Purity : A260/A280 = 1.8-2.0 & A260/A230 = 2.0-2.2

For better result, here are some of our sampling technique recommendations:

  • Use aseptic technique when taking any samples to prevent any unwanted cross-contamination.
  • If the microbial composition data is time-sensitive (e.g samples involving fermentation), extract the genomic DNA immediately after sampling or keep the sample in-20 °C or below.
  • When applicable, consider concentrating any liquid sample with low or sparse abundance using either filtration or centrifugation to improve species detection
  • Raw sequencing data in fastq format.
  • Sequencing data QC.
  • Bioinformatic analysis, includes:
  1. Taxonomy annotation and abundance.
  2. Sankey diagram.
  3. Alpha diversity indices.
  4. Beta diversity statistical analysis (Ordination plot such as NMDS and PCoA).
  5. Various diagrams and charts (Bar chart, Krona, Venn diagram, Heatmap).
  6. Statistical tests (t-test, ANOSIM, MRPP, LEfSe).