Resistance to rust ( Puccinia psidii Winter) in eucalyptus: mode of inheritance and mapping of a major gene with RAPD markers.

Rust is one of the most-damaging eucalypt diseases in Brazil and is considered a potential threat to eucalypt plantations worldwide. To determine the mode of inheritance of resistance in the Eucalyptus grandis- Puccinia psidii pathosystem, ten full-sib families, generated from crosses between susceptible and resistant trees, were inoculated with a single-pustule isolate of the pathogen and rust severity was scored. The observed segregation ratios in segregating families suggested major gene control of rust resistance, although clearly incomplete penetrance, variable expressivity and minor genes are also involved in the global rust-resistance response. To identify markers linked to the resistance locus, screening of RAPD polymorphisms was conducted using bulked segregant analysis in a large full-sib family. A linkage group was built around the Ppr1 gene ( P. psidii resistance gene 1) encompassing six RAPD markers, with a genetic window spanning 5 cM with the two most-closely linked flanking markers. Besides these two flanking markers, RAPD marker AT9/917 co-segregated with Ppr1 without a single recombinant in 994 meioses. This tightly linked marker should prove useful for marker-assisted introgression and will provide an initial lead for a positional cloning effort of this resistance allele. This is the first report of a disease resistance gene identified in Eucalyptus, and one of the few examples of the involvement of a major gene in a non-coevolved pathosystem.

The broad-spectrum tospovirus resistance gene Sw-5 of tomato is a homolog of the root-knot nematode resistance gene Mi.

We used a positional cloning approach to isolate the Sw-5 disease resistance locus of tomato. Complementation experiments with overlapping cosmid clones enabled us to demonstrate that Sw-5 is a single gene locus capable of recognizing several tospovirus isolates and species. Analysis of the predicted Sw-5 protein suggests that it is a cytoplasmic protein, with a potential nucleotide binding site (NBS) domain and a C-terminal end consisting of leucine-rich repeats (LRRs). Based on its structural features, Sw-5 belongs to the class of NBS-LRR resistance genes that includes the tomato Mi, 12, and Prf genes; the Arabidopsis RPM1 gene; and the plant potato virus X resistance gene Rx. The overall similarity between the Sw-5 and Mi proteins of tomato suggests that a shared or comparable signal transduction pathway leads to both virus and nematode resistance in tomato. The similarity also supports the hypothesis that Sw-5 provides resistance via a hypersensitive response. Sw-5 is a member of a loosely clustered gene family in the telomeric region of chromosome 9. Members of this family map to other regions of chromosome 9 and also to chromosome 12, where several fungal, virus, and nematode genes have been mapped, suggesting that paralogs of Sw-5 may have evolved to provide different resistance specificities.

Influence of the antibiotic timentin on plant regeneration of tomato (Lycopersicon esculentum Mill.) cultivars.

Cotyledon explants of tomato (Lycopersicon esculentum Mill. cvs 'Santa Clara', 'Firme' mutant, 'IPA-5' and 'IPA-6') were excised from 8- to 10-day-old in vitro-grown seedlings. Four different shoot induction media supplemented with timentin (300 mg l-1) were screened. When cotyledon explants were cultured on MS-based medium with 1.0 mg l-1 zeatin plus 0.1 mg l-1 IAA and supplemented with timentin, higher regeneration frequencies and a greater number of elongated shoots were obtained. It was observed that timentin caused an increase in the morphogenesis of in vitro cotyledon explants of tomato cultivars. In two of three cultivars tested, rooting of shoots was positively influenced, both in the presence and absence of timentin in the rooting medium, among shoots regenerated from explants derived from timentin-supplemented medium. The results confirm those of a previous investigation on the beneficial effects of this class of antibiotics on tomato regeneration and, consequently, its reliability for use in the transformation of this species.

Hyperhydricity in pepper plants regenerated in vitro: involvement of BiP (Binding Protein) and ultrastructural aspects.

Hyperhydricity in regenerated pepper plants was monitored by the induction of the ER-luminal resident protein, as observed by immunoblotting. Immunoblotting of total protein using an anti-soybean BiP serum indicated that the induction and accumulation of an 80-kDa protein was related to BiP (Binding protein), a 78-kDa ER-resident molecular chaperone. The anti-BiP serum cross-reacted with an 80-kDa protein which was significantly induced by hyperhydricity. Based on similar molecular weight and immunological reactivity we concluded that the 80-kDa protein induced in hyperhydric plants is a BiP homologue. The ultrastructural organisation of leaves in non-hyperhydric and hyperhydric pepper (Capsicum annuum L.) plants was investigated with the aim of identifying the subcellular changes associated with this phenomenon. In non-hyperhydric leaves the chloroplasts of the palisade cells had normally developed thylakoids and grana and a low accumulation or absence of starch grains and plastoglobules. In the hyperhydric plants, however, the chloroplasts exhibited thylakoid disorganisation, low grana number, an accumulation of large starch grains and a low accumulation or absence of plastoglobules. Although the structure of mitochondria and peroxisomes did not change in hyperhydric plants, the number of peroxisomes did increase.

Map-based cloning of the tomato genomic region that spans the Sw-5 tospovirus resistance gene in tomato.

Two yeast artificial chromosomes (YACs) containing genomic DNA from tomato have been isolated using CT220, an RFLP marker which is tightly linked to the tomato spotted wilt virus resistance gene, Sw-5. High-resolution mapping of the YAC ends and internal YAC probes demonstrated that one of the YAC clones, TY257 (400 kb), spans Sw-5. By chromosome walking in a cosmid library, the position of Sw-5 has been delimited within the YAC to a maximal chromosomal segment of 100 kb, spanned by nine overlapping cosmid clones.

Map-based cloning of a protein kinase gene conferring disease resistance in tomato.

The Pto gene in tomato confers resistance to races of Pseudomonas syringae pv. tomato that carry the avirulence gene avrPto. A yeast artificial chromosome clone that spans the Pto region was identified and used to probe a leaf complementary DNA (cDNA) library. A cDNA clone was isolated that represents a gene family, at least six members of which genetically cosegregate with Pto. When susceptible tomato plants were transformed with a cDNA from this family, they were resistant to the pathogen. Analysis of the amino acid sequence revealed similarity to serine-threonine protein kinases, suggesting a role for Pto in a signal transduction pathway.