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Universal ITS2 Barcoding DNA Region Coupled with High-Resolution Melting (HRM) Analysis for Seed Authentication and Adulteration Testing in Leguminous Forage and Pasture Species

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Abstract

Legume species are part of a very important agricultural family, second only to cereals. Their importance for sustainable agriculture worldwide comes from their nitrogen-fixing ability. They include mainly annual grain crops and also very important perennial forage and pasture species. Given their small size, seed admixture and adulteration are a common problem, lowering the forage value, creating weed components in the grassland and causing digestive problems to animals. Here we report the application of the Barcode-DNA High-Resolution Melting (Bar-HRM) analysis method using the universal nuclear plant DNA barcoding region ITS2 for the identification, adulteration and quantification of the main pasture species. Bar-HRM detected Medicago lupulina adulterants in Trifolium pratense seeds as low as 1:100. In conclusion, Bar-HRM analysis could be a faster with higher resolution and cost-effective alternative method to authenticate forage and pasture species and quantitatively detect the purity of their seeds or their feed products.

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Acknowledgments

We thank Dr. Ralli from the Seed Bank of Greece of the National Agricultural Research Foundation for offering the material used in this study, Dr. Argiriou of our institute for his help in preparing the figures of the present manuscript and Mr. Pasentsis for his technical assistance. We also thank Mrs. Despoina Loukidou and Mrs. Laura Dadurian for proofreading the manuscript. The continuous support of the General Secretariat for Research and Technology is also acknowledged.

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Authors and Affiliations

  1. Department of Genetics and Plant Breeding, School of Agriculture, Aristotle University of Thessaloniki, Thessaloniki, 54 124, Greece

    Ioannis Ganopoulos & Athanasios Tsaftaris

  2. Institute of Agrobiotechnology, CERTH, 6th km Charilaou-Thermis Road, Thermi, Thessaloniki, 57001, Greece

    Ioannis Ganopoulos, Panagiotis Madesis & Athanasios Tsaftaris

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  1. Ioannis Ganopoulos
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  2. Panagiotis Madesis
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  3. Athanasios Tsaftaris
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Correspondence to Athanasios Tsaftaris.

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Ioannis Ganopoulos and Panagiotis Madesis contributed equally for this work.

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Ganopoulos, I., Madesis, P. & Tsaftaris, A. Universal ITS2 Barcoding DNA Region Coupled with High-Resolution Melting (HRM) Analysis for Seed Authentication and Adulteration Testing in Leguminous Forage and Pasture Species. Plant Mol Biol Rep 30, 1322–1328 (2012). https://doi.org/10.1007/s11105-012-0453-3

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