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Characterization of novel wheat NBS domain-containing sequences and their utilization, in silico, for genome-scale R-gene mining

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Abstract

In crop improvement, the isolation, cloning and transfer of disease resistance genes (R-genes) is an ultimate goal usually starting from tentative R-gene analogs (RGAs) that are identified on the basis of their structure. For bread wheat, recent advances in genome sequencing are supporting the efforts of wheat geneticists worldwide. Among wheat R-genes, nucleotide-binding site (NBS)-encoding ones represent a major class. In this study, we have used a polymerase chain reaction-based approach to amplify and clone NBS-type RGAs from a bread wheat cultivar, ‘Salambo 80.’ Four novel complete ORF sequences showing similarities to previously reported R-genes/RGAs were used for in silico analyses. In a first step, where analyses were focused on the NBS domain, these sequences were phylogenetically assigned to two distinct groups: a first group close to leaf rust Lr21 resistance proteins; and a second one similar to cyst nematode resistance proteins. In a second step, sequences were used as initial seeds to walk up and downstream the NBS domain. This procedure enabled identifying 8 loci ranging in size between 2,115 and 7,653 bp. Ab initio gene prediction identified 8 gene models, among which two had complete ORFs. While GenBank survey confirmed the belonging of sequences to two groups, subsequent characterization using IWGSC genomic and proteomic data showed that the 8 gene models, reported in this study, were unique and their loci matched scaffolds on chromosome arms 1AS, 1BS, 4BS and 1DS. The gene model located on 1DS is a pseudo-Lr21 that was shown to have an NBS-LRR domain structure, while the potential association of the RGAs, here reported, is discussed. This study has produced novel R-gene-like loci and models in the wheat genome and provides the first steps toward further elucidation of their role in wheat disease resistance.

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Acknowledgments

We gratefully acknowledge Abdennour Sébéi (Centre Régional de Recherches sur les Grandes Cultures, Béja, Tunisia) for providing bread wheat cultivar ‘Salambo 80’; Jacques-Deric Rouault (Laboratoire Evolution, Génome et Spéciation, CNRS, Gif sur Yvette, France) for providing training on data mining techniques; Paul A. Wilkinson (School of Biological Sciences, University of Bristol, UK) for providing help about CerealsDB 2.0.; and Abdelkader Aïnouche (UMR-CNRS Ecobio, Université de Rennes-1, France) for reading the manuscript. This study was financially supported by the Tunisian Ministry of Higher Education and Scientific Research.

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Authors and Affiliations

  1. Unité de Recherche Génomique des Insectes Ravageurs des Cultures d’intérêt agronomique (GIRC, UR11ES10), Faculté des Sciences de Tunis, Université de Tunis El-Manar, El-Manar, 2092, Tunis, Tunisia

    Dhia Bouktila, Yosra Habachi-Houimli, Yosra Khalfallah, Maha Mezghani-Khemakhem, Mohamed Makni & Hanem Makni

  2. Institut Supérieur de Biotechnologie de Béja (ISBB), Université de Jendouba, 9000, Béja, Tunisia

    Dhia Bouktila

  3. Institut Supérieur de l’Animation pour la Jeunesse et la Culture (ISAJC), Université de Tunis, Bir-El-Bey, Tunisia

    Hanem Makni

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  1. Dhia Bouktila
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  2. Yosra Habachi-Houimli
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  3. Yosra Khalfallah
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  4. Maha Mezghani-Khemakhem
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  5. Mohamed Makni
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  6. Hanem Makni
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Corresponding author

Correspondence to Dhia Bouktila.

Additional information

Communicated by S. Hohmann.

Nucleotide sequence data reported are available in the GenBank database under the accession numbers JX566982 to JX566985.

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Bouktila, D., Habachi-Houimli, Y., Khalfallah, Y. et al. Characterization of novel wheat NBS domain-containing sequences and their utilization, in silico, for genome-scale R-gene mining. Mol Genet Genomics 289, 599–613 (2014). https://doi.org/10.1007/s00438-014-0834-4

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