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HRM technology for the identification and characterization of INDEL and SNPs mutations in genes involved in drought and salt tolerance of durum wheat

Published online by Cambridge University Press:  16 March 2011

Linda Mondini ,
Miloudi M. Nachit ,
Enrico Porceddu  and
Mario A. Pagnotta
Linda Mondini
Affiliation:
Department of Agrobiology and Agrochemistry, University of Tuscia, Via S.C. de Lellis, 01100 Viterbo, Italy
Miloudi M. Nachit
Affiliation:
ICARDA, PO Box 5466, Aleppo, Syria
Enrico Porceddu
Affiliation:
Department of Agrobiology and Agrochemistry, University of Tuscia, Via S.C. de Lellis, 01100 Viterbo, Italy
Mario A. Pagnotta*
Affiliation:
Department of Agrobiology and Agrochemistry, University of Tuscia, Via S.C. de Lellis, 01100 Viterbo, Italy
*
*Corresponding author. E-mail: [email protected]
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Abstract

WRKY transcription factors are one of the largest families of transcriptional regulators and form an integral part of signalling webs which modulate many plant processes, such as abiotic stress tolerance. In the present paper, an innovative method has been applied to identify novel WRKY-1 alleles involved in the responses to salt and drought stresses in Triticum durum. This technique involves scanning for sequencing variations in cDNA-derived PCR amplicons, using high-resolution melting (HRM) followed by direct Sanger sequencing of only those amplicons which were predicted to carry nucleotide changes. HRM represents a novel advance in detection of single-nucleotide polymorphisms (SNPs) by measuring temperature-induced strand separation of short PCR amplicons. The use of this approach is still limited in the field of plant biology. Here, HRM analysis has been applied to the discovery and genotyping of durum wheat SNPs. Specific primers have been designed, starting at multi-alignment of WRKY-1-conserved portions. The PCR amplicons, containing single SNPs, produce distinctive HRM profiles, and by sequencing the PCR products identified, SNPs have been characterized and validated. The results showed that all the revealed SNPs are located on salt-tolerant varieties, confirming their value in breeding activities.

Keywords

durum wheathigh-resolution meltingsalt tolerancesingle-nucleotide polymorphisms
Type
Research Article
Information
Plant Genetic Resources , Volume 9 , Issue 2 , July 2011 , pp. 166 - 169
Copyright
Copyright © NIAB 2011

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