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Appl Plant Sci. 2018 Dec 11;6(12):e01201. doi: 10.1002/aps3.1201. eCollection 2018 Dec.

Advances in genotyping microsatellite markers through sequencing and consequences of scoring methods for Ceratonia siliqua (Leguminosae).

Author information

Royal Botanic Gardens Kew, Richmond Surrey TW9 3DS United Kingdom.
Institut Méditerranéen de Biodiversité et d'Ecologie marine et continentale (IMBE) [IMBE is sponsored by Aix Marseille University, Avignon University, Centre National de la Recherche Scientifique (CNRS), and Institut de Recherche pour le Développement (IRD)] Station marine d'Endoume, Chemin de la Batterie des Lions FR-13007 Marseille France.
Institut du Cerveau et de la Moelle épinière (ICM) Hôpital Pitié Salpêtrière 47 Boulevard de l'Hôpital 75013 Paris France.
Laboratoire Caractérisation Génétique des Plantes Faculté des sciences Université Saint-Joseph B.P. 11-514 Riad El Solh Beirut 1107 2050 Lebanon.
Laboratoire d'Ecologie et Environnement Faculté des Sciences Semlalia Université Cadi Ayyad Marrakesh Morocco.
Dipartimento di Agricoltura, Alimentazione e Ambiente (Di3A) Università degli Studi di Catania Via Valdisavoia 5 95123 Catania Italy.
Centre de coopération internationale en recherche agronomique pour le développement (CIRAD) Laboratoire des Symbioses Tropicales et Méditerranéennes (LSTM) Montpellier France.
LSTM [LSTM is sponsored by University of Montpellier, CIRAD, IRD, INRA, Montpellier SupAgro] TA A-82/J Campus International de Baillarguet FR-34398 Montpellier CEDEX 5 France.
Real Jardín Botánico (CSIC) Plaza de Murillo 2 28014 Madrid Spain.


Premise of the Study:

Simple sequence repeat (SSR) or microsatellite markers have been used in a broad range of studies mostly scoring alleles on the basis of amplicon size as a proxy for the number of repeat units of an SSR motif. However, additional sources of variation within the SSR or in the flanking regions have largely remained undetected.


In this study, we implemented a next-generation sequencing-based genotyping approach in a newly characterized set of 18 nuclear SSR markers for the carob tree, Ceratonia siliqua. Our aim was to evaluate the effect of three different methods of scoring molecular variation present within microsatellite markers on the genetic diversity and structure results.


The analysis of the sequences of 77 multilocus genotypes from four populations revealed SSR variation and additional sources of polymorphism in 87% of the loci analyzed (42 single-nucleotide polymorphisms and five insertion/deletion polymorphisms), as well as divergent paralog copies in two loci. Ignoring sequence variation under standard amplicon size genotyping resulted in incorrect identification of 69% of the alleles, with important effects on the genetic diversity and structure estimates.


Next-generation sequencing allows the detection and scoring of SSRs, single-nucleotide polymorphisms, and insertion/deletion polymorphisms to increase the resolution of population genetic studies.


MicNeSs; carob tree; genetic diversity; homoplasy; next‐generation sequencing; simple sequence repeat (SSR)

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