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Assessment of high-resolution melting (HRM) profiles as predictors of microsatellite variation: an example in Providence Petrel (Pterodroma solandri)

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Abstract

High-resolution melting (HRM) analysis is an emerging technology to screen microsatellites for polymorphism. A potential issue surrounding this method is that amplicon sizes for HRM should typically be short (80–100 bp) for highest sensitivity to reveal polymorphism via the presence of two peaks in the curve of the derivative of fluorescence over temperature (dF/dT). In contrast, microsatellite amplicons are typically 100–400 bp. Therefore, we compared HRM analysis melting temperature range (ΔTm) and multiple dF/dT peaks for predicting microsatellite polymorphism. We assessed polymorphism at 27 microsatellite loci, with estimated lengths of 122–321 bp, in Providence Petrel (Pterodroma solandri). We validated HRM assessment using traditional capillary electrophoresis (CE). While 100 % of loci exhibiting multiple peaks in the dF/dT curve were confirmed as polymorphic by CE, 16 % improvement in sensitivity (83 vs. 67 %) was achieved by using ΔTm, and 25 % (92 vs. 67 %) by using ΔTm in addition to multiple dF/dT peaks. We suggest HRM melting temperature range as new predictor of polymorphism that can be used to rapidly assess microsatellites polymorphism.

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Acknowledgements

The Seaworld Research and Rescue Foundation Inc (Grant SWR/4/2011) supported this work.

Author contributions

Anicee Lombal and Theodore Wenner constructed the manuscript and collected the molecular data. Anicee Lombal performed statistical analyses. Christopher Burridge designed the study and contributed to the manuscript.

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Authors and Affiliations

  1. School of Biological Sciences, University of Tasmania, Private Bag 55, Hobart, TAS, 7001, Australia

    Anicee J. Lombal, Theodore J. Wenner & Christopher P. Burridge

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  1. Anicee J. Lombal
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  2. Theodore J. Wenner
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  3. Christopher P. Burridge
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Correspondence to Anicee J. Lombal.

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Lombal, A.J., Wenner, T.J. & Burridge, C.P. Assessment of high-resolution melting (HRM) profiles as predictors of microsatellite variation: an example in Providence Petrel (Pterodroma solandri). Genes Genom 37, 977–983 (2015). https://doi.org/10.1007/s13258-015-0327-9

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  • DOI: https://doi.org/10.1007/s13258-015-0327-9

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