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Mol Ecol Resour. 2011 Mar;11(2):394-9. doi: 10.1111/j.1755-0998.2010.02933.x. Epub 2010 Nov 2.

The use of high-resolution melting analysis for genotyping Symbiodinium strains: a sensitive and fast approach.

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1
Department of Biology, University of Louisiana at Lafayette, Lafayette, LA 70504, USA.

Abstract

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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