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Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons
Published on: August 29, 2014
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Detecting invertebrate species in archived collections using next-generation sequencing
Melissa E Carew1, Leon Metzeling2, Rosalind St Clair2
1School of BioSciences, Bio21 Institute, Victorian Centre for Aquatic Pollution Identification and Management (CAPIM), The University of Melbourne, 30 Flemington Rd, Melbourne, VIC, 3010, Australia.
Molecular Ecology Resources
|December 18, 2016
Summary
Next-generation sequencing (NGS) shows potential for identifying species in archived aquatic macroinvertebrate samples. While effective for common taxa, older samples and gaps in DNA barcode libraries present challenges for comprehensive biomonitoring.
Area of Science:
- Environmental Science
- Molecular Biology
- Ecology
Background:
- Traditional biological monitoring relies on family-level invertebrate biodiversity to assess ecosystem health.
- Next-generation sequencing (NGS) offers potential for enhanced species identification, improving biomonitoring sensitivity.
- Utilizing archived collections with environmental data can aid in understanding species distributions and responses.
Purpose of the Study:
- To evaluate the efficacy of NGS for detecting species in archived macroinvertebrate samples.
- To assess the impact of sample age and DNA barcode library completeness on NGS-based species identification.
Main Methods:
- Archived macroinvertebrate samples (stored up to 12 years in ethanol) were analyzed using NGS.
- Three DNA barcode amplicons (197–274 bp) were amplified and compared against existing DNA barcoding libraries.
- Species identification success was assessed based on amplicon detection and library matching.
Main Results:
- Partial DNA barcodes were amplified from most samples, enabling species detection, often with multiple amplicons.
- Older samples (>8 years) yielded fewer taxa and less reliable amplification compared to younger samples.
- Common taxa with existing DNA barcodes were reliably identified, demonstrating NGS potential for archived material.
Conclusions:
- NGS can successfully identify many common aquatic macroinvertebrate species in archived samples, aiding long-term biomonitoring.
- Gaps in DNA barcode libraries and sample degradation in older specimens (over 8 years) limit comprehensive species detection.
- Further development of DNA barcode libraries and optimized protocols for older samples are needed for broader application.
