16S rRNA Sequencing Technology
What is 16S rRNA Sequencing?
16S rRNA sequencing is a targeted DNA sequencing method used to identify and classify bacteria. It sequences a specific gene (16S ribosomal RNA gene) that is present in all bacteria but has variable regions unique to different species.
Why Use 16S rRNA?
The 16S rRNA gene is ideal for bacterial identification because:
- Universal: Present in all bacteria and archaea
- Conserved Regions: Allow universal primers to amplify
- Variable Regions: Enable species-level identification
- Well-Characterized: Extensive reference databases exist
How It Works
- DNA Extraction: Extract DNA from sample
- PCR Amplification: Amplify 16S rRNA gene using universal primers
- Sequencing: Sequence the amplified regions (V1-V9 variable regions)
- Bioinformatics: Compare sequences to reference databases
Variable Regions
The 16S gene has 9 variable regions (V1-V9):
- V3-V4: Most commonly sequenced; good resolution
- V1-V2: Good for certain bacterial groups
- Full-length: Highest resolution but more expensive
Advantages
- Cost-effective: Cheaper than shotgun metagenomics
- Well-established: Standardized protocols and databases
- Focused: Specifically identifies bacteria
Limitations
- No Functional Info: Only identifies organisms, not their genes
- PCR Bias: Some bacteria may amplify better than others
- Limited Resolution: May not distinguish closely related species
Related Terms
- Metagenomics: Broader sequencing approach
- WGS: Whole Genome Sequencing
- Microbiome: The community being studied
- Alpha Diversity: Diversity calculated from 16S data