Hostname: page-component-7c8c6479df-7qhmt Total loading time: 0 Render date: 2024-03-27T19:13:35.024Z Has data issue: false hasContentIssue false

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
Affiliation:
Department of Agrobiology and Agrochemistry, University of Tuscia, Via S.C. de Lellis, 01100Viterbo, 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, 01100Viterbo, Italy
Mario A. Pagnotta*
Affiliation:
Department of Agrobiology and Agrochemistry, University of Tuscia, Via S.C. de Lellis, 01100Viterbo, Italy
*
*Corresponding author. E-mail: pagnotta@unitus.it

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.

Type
Research Article
Copyright
Copyright © NIAB 2011

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Boyer, JS (1982) Plant productivity and environment. Science 218: 443448.CrossRefGoogle ScholarPubMed
Chagné, D, Carlisle, CM, Blond, C, Volz, RK, Whitworth, CJ, Oraguzie, NC, Crowhurst, RN, Allan, AC, Espley, RV, Hellens, RP and Gardiner, SE (2007) Mapping a candidate gene (MdMYB10) for red flesh and foliage colour in apple. BMC Genomics 8: 212.CrossRefGoogle ScholarPubMed
Croxford, AE, Rogers, T, Caligari, PDS and Whilkinson, MJ (2008) High-resolution melt analysis to identify and map sequence-tagged site anchor points onto linkage maps: a white lupin (Lupinus albus) map as an exemplar. New Phytologist 180: 594607.CrossRefGoogle Scholar
Hasegawa, PM and Bressan, RE (2000) Plant cellular and molecular responses to high salinity. Annual Review of Plant Physiology and Plant Molecular Biology 51: 463499.CrossRefGoogle ScholarPubMed
Ishiguro, S and Nakamura, K (1994) Characterization of a cDNA encoding a novel DNA-binding protein, SPF1, that recognizes SP8 sequences in the 5′ upstream regions of genes coding for sporamin and beta-amylase from sweet potato. Molecular and General Genetics 244: 563571.CrossRefGoogle ScholarPubMed
Kader, MA, Seidel, T, Golldack, D and Lindberg, S (2006) Expressions of OsHKT1, OsHKT2, and OsVHA are differentially regulated under NaCl stress in salt-sensitive and salt-tolerant rice (Oryza sativa L.) cultivars. Journal of Experimental Botany 57: 42574268.CrossRefGoogle ScholarPubMed
Kennerson, L, Warburton, T, Nelis, E, Brewer, M, Polly, P, De Jonghe, P, Timmerman, V and Nicholson, GA (2007) Mutation scanning the GJB1 gene with high-resolution melting analysis: implication for mutation scanning of genes for Charcot-Marie-Tooth disease. Clinical Chemistry 53: 349352.CrossRefGoogle ScholarPubMed
Rushton, PJ, Torres, JT, Parniske, M, Wernert, P, Hahlbrock, K and Somssich, IE (1996) Interaction of elicitor-induced DNA binding proteins with elicitor response elements in the promoters of parsley PR1 genes. EMBO Journal 15: 56905700.CrossRefGoogle ScholarPubMed
Rushton, PJ, Somssich, IE, Ringler, P and Shen, QJ (2010) WRKY transcription factors. Trends in Plant Science: 15: 247258.Google ScholarPubMed