Proceedings of the XLVI Italian Society of Agricultural Genetics - SIGA Annual Congress

Giardini Naxos, Italy - 18/21 September, 2002

ISBN 88-900622-3-1

Poster Abstract - 4.31

PROMOTER ANALYSIS OF PATHOGENESIS-RELATED GENES IN RESISTANT AND SUSCEPTIBLE TOMATO CULTIVARS AND LYCOPERSICON WILD SPECIES

SORACE S.*, GORI M.**, BETTINI P.*

*) Dipartimento di Biologia animale e Genetica “Leo Pardi”, Università degli Studi di Firenze

**) Centro Interdipartimentale di servizi per le Biotecnologie di Interesse Agrario, Chimico e Industriale, Università degli Studi di Firenze (C.I.B.I.A.C.I.)

tomato, plant-pathogen interaction, PR-proteins, non-coding sequences

Genomes are known to contain coding, but also non-coding, sequences, whose relative abundance has been varying throughout evolution. Non-coding sequences have been considered for a long time "junk DNA" devoid of any biological meaning, but the discovery of a large array of regulatory functions has changed this view, and led to a new attention to the features of upstream, intervening and downstream DNA tracts. DNA secondary structures (hairpins, cruciforms, triple helixes), whose formation depends on specific homogeneous sequences, have been correlated with biological activity and gene regulation in many instances (Catasti et al. 1999, Genetica 106: 15-36; Catasti et al. 1996, J. Mol. Biol. 264: 534-545; Iyer et al. 1995, EMBO J. 14: 2570-2579).

Here we report the first part of a study on the correlation between variations in non-coding DNA sequences, i.e. promoters, and the level of gene expression in a group of genes coding for pathogenesis-related (PR) proteins, known to play a crucial role in host-pathogen interaction both at the local and the systemic level, with the aim of developing markers for tolerance/resistance to disease.

Promoters of PR-genes have been shown to contain sequences involved in the induction of gene expression upon wounding and after treatment with ethylene, methyl jasmonate, salicylic acid or other elicitors. In particular, the so-called PR box or GCC box is found in many plant defense-related genes promoters, including the promoters of many basic PR proteins. Based on these data, we have chosen two genes whose promoters contain one or more GCC boxes, i.e. endochitinase and osmotin (Jia Y and Martin GB 1999, Plant Molecular Biology 40: 455-465). Moreover, as PR-1 gene is considered to be a marker for Systemic Acquired Resistance (SAR), two PR-1 genes differentially regulated during the hypersensitive response and development have been included (Tornero P et al. 1997, Mol. Plant Microbe Interact. 10: 624-634): PR1a2, whose expression is costitutive, and PR1b1 that is induced by pathogen attack, salicylic acid and ethylene precursors.

The study has been carried out on 14 Lycopersicon esculentum cultivars known to be susceptible or resistant to pathogens, as well as on the wild species L. pennellii, L. pimpinellifolium and L. peruvianum.

Primers for PCR amplification have been designed on the promoter sequences found in GenBank, the reverse primer being anchored in the coding region to increase specificity. Amplified sequences have been cloned, sequenced and aligned in order to detect the presence of polymorphisms.

Preliminary data show that both single nucleotide polymorphisms (SNPs) and variations in microsatellite length can be detected in the promoters of the above mentioned genes, the sequences of the wild species showing the highest variability.

Research supported by the Ministero delle Politiche Agricole e Forestali (MiPAF) in the frame of the coordinated project "Plant protection through the use of molecular markers" (PROMAR).