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.     

Bayesian and maximum likelihood estimation of genetic maps

There has recently been increased interest in the use of Markov Chain Monte Carlo (MCMC)-based Bayesian methods for estimating genetic maps. The advantage of these methods is that they can deal accurately with missing data and genotyping errors. Here we present an extension of the previous methods that makes the Bayesian method applicable to large data sets. We present an extensive simulation study examining the statistical properties of the method and comparing it with the likelihood method implemented in Mapmaker. We show that the Maximum A Posteriori (MAP) estimator of the genetic distances, corresponding to the maximum likelihood estimator, performs better than estimators based on the posterior expectation. We also show that while the performance is similar between Mapmaker and the MCMC-based method in the absence of genotyping errors, the MCMC-based method has a distinct advantage in the presence of genotyping errors. A similar advantage of the Bayesian method was not observed for missing data. We also re-analyse a recently published set of data from the eggplant and show that the use of the MCMC-based method leads to smaller estimates of genetic distances.     

QTL analysis of an advanced backcross of Lycopersicon peruvianum to the cultivated tomato and comparisons with QTLs found in other wild species

 A BC₃ population previously developed from a backcross of Lycopersicon peruvianum, a wild relative of tomato, into the cultivated variety L. esculentum was analyzed for QTLs. Approximately 200 BC₄ families were scored for 35 traits in four locations worldwide. One hundred and sixty-six QTLs were detected for 29 of those traits. For more than half of those 29 traits at least 1 QTL was detected for which the presence of the wild allele was associated with an agronomically beneficial effect despite the inferior phenotype of the wild parent. Eight QTLs for fruit weight could be followed through the BC₂, BC₃, and BC₄, generations, supporting the authenticity of these QTLs. Comparisons were made between the QTLs found in this study and those found in studies involving two other wild species; the results showed that while some of these QTLs can be presumed to be allelic, most of the QTLs detected in this study are ones not previously discovered. Received: 9 April 1997 / Accepted: 20 May 1997     

Molecular mapping of rice chromosomes

Summary We report the construction of an RFLP genetic map of rice (Oryza sativa) chromosomes. The map is comprised of 135 loci corresponding to clones selected from a PstI genomic library. This molecular map covers 1,389 cM of the rice genome and exceeds the current classical maps by more than 20%. The map was generated from F₂ segregation data (50 individuals) from a cross between an indica and javanica rice cultivar. Primary trisomics were used to assign linkage groups to each of the 12 rice chromosomes. Seventy-eight percent of the clones assayed revealed RFLPs between the two parental cultivars, indicating that rice contains a significant amount of RFLP variation. Strong correlations between size of hybridizing restriction fragments and level of polymorphism indicate that a significant proportion of the RFLPs in rice are generated by insertions/delections. This conclusion is supported by the occurrence of null alleles for some clones (presumably created by insertion or deletion events). One clone, RG229, hybridized to sequences in both the indica and javanica genomes, which have apparently transposed since the divergence of the two cultivars from their last common ancestor, providing evidence for sequence movement in rice. As a by product of this mapping project, we have discovered that rice DNA is less C-methylated than tomato or maize DNA. Our results also suggest the notion that a large fraction of the rice genome (approximately 50%) is single copy.     

CR1 — a dispersed repeated element associated with the Cab-1 locus in tomato

Cab-1 is a complex genetic locus in tomato consisting of four clustered genes encoding chlorophyll a/b-binding polypeptide. Southern blot analysis of total tomato DNA with genomic clones corresponding to the Cab-1 locus has revealed the presence of a repetitive element in the 3 kb spacer regions between two of these genes. This repetitive element, named CR1, has been characterized via sequencing, genetic mapping and hybridization to related solanaceous species. Results indicate that there are as many as 30 copies of this element in the tomato genome and that most, if not all, are found at independent loci. Sites corresponding to 12 of the repeats have been located on different regions of chromosomes 2, 4, 5, 7, 10 and 11. A 1.6 kb PstI-EcoRI fragment from the Cab-1 locus containing the element was sequenced and found to be 75% AT-rich. No open reading frames larger than 150 bp were detected. Several imperfect inverted repeats flanked by direct repeats could be found at the ends of the element. This arrangement is reminiscent of known transposons. Southern hybridization analysis indicates that multiple copies of CR1 exist in all species of the genus Lycopersicon as well as in Solanum lycopersicoides and S. tuberosum (potato), but not in eggplant, pepper, petunia, Datura or tobacco. Melt-off experiments indicate that members of the CR1 family in the tomato genome are more closely related to one another than to homologous members in the genomes of S. lycopersicoides or S. tuberosum, suggesting some type of concerted evolution.     

Advanced backcross QTL analysis of tomato. II. Evaluation of near-isogenic lines carrying single-donor introgressions for desirable wild QTL-alleles derived from Lycopersicon hirsutum and L. pimpinellifolium

 Improved-processing tomato lines were produced by the molecular breeding strategy of advanced backcross QTL (AB-QTL) analysis. These near-isogenic lines (NILs) contained unique introgressions of wild alleles originating from two donor wild species, Lycopersicon hirsutum (LA1777) and L. pimpinellifolium (LA1589). Wild alleles targeted for trait improvement were selected on the basis of previously published replicated QTL data obtained from advanced backcross populations for a battery of important agronomic traits. Twenty three NILs were developed for 15 genomic regions which were predicted to contain 25 quantitative trait factors for the improvement of seven agronomic traits: total yield, red yield, soluble solids, brix×red yield, viscosity, fruit color, and fruit firmness. An evaluation of the agronomic performance of the NILs in five locations worldwide revealed that 22 out of the 25 (88%) quantitative factors showed the phenotypic improvement predicted by QTL analysis of the BC₃ populations, as NILs in at least one location. Per-location gains over the elite control ranged from 9% to 59% for brix×red yield; 14% to 33% for fruit color; 17% to 34% for fruit firmness; 6% to 22% for soluble-solids content; 7% to 22% for viscosity; 15% to 48% for red yield, and 20% to 28% for total yield. The inheritance of QTLs, the implementation of the AB-QTL methodology for characterizing unadapted germplasm and the applicability of this method to other crops are discussed. Received: 27 October 1997 / Accepted: 25 November 1997     

Advanced backcross QTL analysis in tomato. I. Identification of QTLs for traits of agronomic importance from Lycopersicon hirsutum

 Advanced backcross QTL (AB-QTL) analysis is a new strategy for studying the effect of unadapted alleles on the agronomic performance of elite cultivated lines. In this paper we report results from the application of the AB-QTL strategy to cultivated tomato using the wild species Lycopersicon hirsutum LA1777 as the donor parent. RFLP genomic fingerprints were determined for 315 BC₂ plants and phenotypic data were collected for 19 agronomic traits from approximately 200 derived BC₃ lines which were grown in replicated field trials in three locations worldwide. Between 1 and 12 significant QTLs were identified for each of the 19 traits evaluated, with a total of 121 QTLs identified for all traits. For 25 of the QTLs (20%) corresponding to 12 traits (60%), the L. hirsutum allele was associated with an improvement of the trait from a horticultural perspective, despite the fact that L. hirsutum is overall phenotypically inferior to the elite parent. For example, L. hirsutum has fruit that remains green when ripe (lack of red pigment) yet alleles were found in this species that significantly increase red color when transferred into cultivated tomatoes. Wild alleles were also associated with increases in total yield and soluble solids (up to 15%) and brix×red yield (up to 41%). These results support the idea that one cannot predict the genetic potential of exotic germplasm based on phenotype alone and that marker-based methods, such as the AB-QTL strategy, should be applied to fully exploit exotic germplasm. Received: 27 October 1997 / Accepted: 25 November 1997     

The tomato Never-ripe locus regulates ethylene-inducible gene expression and is linked to a homolog of the Arabidopsis ETR1 gene.

Fruit ripening represents a complex system of genetic and hormonal regulation of eukaryotic development unique to plants. We are using tomato ripening mutants as tools to elucidate genetic components of ripening regulation and have recently demonstrated that the Never-ripe (Nr) mutant is insensitive to the plant growth regulator ethylene (M.B. Lanahan, H.-C. Yen, J.J. Giovannoni, H.J. Klee [1994] Plant Cell 6:521-530). We report here ethylene sensitivity over a range of concentrations in normal and Nr tomato seedlings and show that the Nr mutant retains residual sensitivity to as little as 1 part per million of ethylene. Analysis of ripening-related gene expression in normal and mutant ethylene-treated fruit demonstrates that Nr exerts its influence on development at least in part at the level of ethylene-inducible gene expression. We have additionally used cloned tomato and Arabidopsis sequences known to influence ethylene perception as restriction fragment length polymorphism probes, and have identified a tomato locus linked to Nr that hybridizes to the Arabidopsis ETR1 gene at low stringency, suggesting the possibility that Nr may be homologous to ETR1.     

Recombinant inbred lines for genetic mapping in tomato

A cross between the cultivated tomato Lycopersicon esculentum and a related wild species L. cheesmanii yielded 97 recombinant inbred lines (RILs) which were used to construct a genetic map consisting of 132 molecular markers. Significant deviation from the expected 1:1 ratio between the two homozygous classes was found in 73% of the markers. In 98% of the deviating markers, L. esculentum alleles were present in greater frequency than the L. cheesmanii alleles. For most of the markers with skewed segregation, the direction of the deviation was maintained from F₂ to F₇ generations. The average heterozygosity in the population was 15%. This value is significantly greater than the 1.5% heterozygosity expected for RILs in the F₇ generation. On average, recombination between linked markers was twice as high in the RILs than in the F₂ population used to derive them. The utility of RILs for the mapping of qualitative and quantitative traits is discussed.     

Abundance, variability and chromosomal location of microsatellites in wheat

The potential of microsatellite sequences as genetic markers in hexaploid wheat (Triticum aestivum) was investigated with respect to their abundance, variability, chromosomal location and usefulness in related species. By screening a lambda phage library, the total number of (GA)_n blocks was estimated to be 3.6 x 10⁴ and the number of (GT)_n blocks to be 2.3 x 10⁴ per haploid wheat genome. This results in an average distance of approximately 270 kb between these two microsatellite types combined. Based on sequence analysis data from 70 isolated microsatellites, it was found that wheat microsatellites are relatively long containing up to 40 dinucleotide repeats. Of the tested primer pairs, 36% resulted in fragments with a size corresponding to the expected length of the sequenced microsatellite clone. The variability of 15 microsatellite markers was investigated on 18 wheat accessions. Significantly, more variation was detected with the microsatellite markers than with RFLP markers with, on average, 4.6 different alleles per microsatellite. The 15 PCR-amplified microsatellites were further localized on chromosome arms using cytogenetic stocks of Chinese Spring. Finally, the primers for the 15 wheat microsatellites were used for PCR amplification with rye (Secale cereale) and barley accessions (Hordeum vulgare, H. spontaneum). Amplified fragments were observed for ten primer pairs with barley DNA and for nine primer pairs with rye DNA as template. A microsatellite was found by dot blot analysis in the PCR products of barley and rye DNA for only one primer pair.