Journal
MOLECULAR ECOLOGY RESOURCES
Volume 11, Issue 2, Pages 394-399Publisher
WILEY
DOI: 10.1111/j.1755-0998.2010.02933.x
Keywords
corals; denaturing gradient gel electrophoresis; genotyping; high-resolution melting; ITS2; Symbiodinium; symbiotic dinoflagellates
Funding
- NSF-OCE [0851123]
- PADI Foundation
- Directorate For Geosciences
- Division Of Ocean Sciences [1321500] Funding Source: National Science Foundation
- Directorate For Geosciences
- Division Of Ocean Sciences [0851123] Funding Source: National Science Foundation
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High-resolution melting (HRM) analysis is a closed-tube, rapid and sensitive technique able to detect DNA variations. It relies on the fluorescence melting curves that are obtained from the transition of double-stranded DNA (dsDNA) to single-stranded DNA (ssDNA) as a result of temperature increase. In this study, we evaluated the effectiveness of HRM as a tool to rapidly and precisely genotype monotypic Symbiodinium populations using the internal transcribed spacer, region 2, ribosomal DNA (ITS2 rDNA). For this, Symbiodinium denaturing gradient gel electrophoresis (DGGE) profiles, where gel bands were excised and sequenced, were compared to HRM genotypes. Results showed that twenty cultures were correctly genotyped in < 2 h using HRM analysis with a percentage of confidence > 90%. Limitations of the technique were also investigated. Unlike other techniques used for genotyping Symbiodinium, such as DGGE and other fingerprint profiles, HRM is a technique of great advantage for field coral reef ecologists and physiologists as no expertise in advanced molecular methods is required.
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