Analysis of expressed sequence tags generated from full-length enriched cDNA libraries of melon.

BACKGROUND: Melon (Cucumis melo), an economically important vegetable crop, belongs to the Cucurbitaceae family which includes several other important crops such as watermelon, cucumber, and pumpkin. It has served as a model system for sex determination and vascular biology studies. However, genomic resources currently available for melon are limited. RESULT: We constructed eleven full-length enriched and four standard cDNA libraries from fruits, flowers, leaves, roots, cotyledons, and calluses of four different melon genotypes, and generated 71,577 and 22,179 ESTs from full-length enriched and standard cDNA libraries, respectively. These ESTs, together with ~35,000 ESTs available in public domains, were assembled into 24,444 unigenes, which were extensively annotated by comparing their sequences to different protein and functional domain databases, assigning them Gene Ontology (GO) terms, and mapping them onto metabolic pathways. Comparative analysis of melon unigenes and other plant genomes revealed that 75% to 85% of melon unigenes had homologs in other dicot plants, while approximately 70% had homologs in monocot plants. The analysis also identified 6,972 gene families that were conserved across dicot and monocot plants, and 181, 1,192, and 220 gene families specific to fleshy fruit-bearing plants, the Cucurbitaceae family, and melon, respectively. Digital expression analysis identified a total of 175 tissue-specific genes, which provides a valuable gene sequence resource for future genomics and functional studies. Furthermore, we identified 4,068 simple sequence repeats (SSRs) and 3,073 single nucleotide polymorphisms (SNPs) in the melon EST collection. Finally, we obtained a total of 1,382 melon full-length transcripts through the analysis of full-length enriched cDNA clones that were sequenced from both ends. Analysis of these full-length transcripts indicated that sizes of melon 5' and 3' UTRs were similar to those of tomato, but longer than many other dicot plants. Codon usages of melon full-length transcripts were largely similar to those of Arabidopsis coding sequences. CONCLUSION: The collection of melon ESTs generated from full-length enriched and standard cDNA libraries is expected to play significant roles in annotating the melon genome. The ESTs and associated analysis results will be useful resources for gene discovery, functional analysis, marker-assisted breeding of melon and closely related species, comparative genomic studies and for gaining insights into gene expression patterns.

Mapping and linkage disequilibrium analysis with a genome-wide collection of SNPs that detect polymorphism in cultivated tomato.

The history of tomato (Solanum lycopersicum L.) improvement includes genetic bottlenecks, wild species introgressions, and divergence into distinct market classes. This history makes tomato an excellent model to investigate the effects of selection on genome variation. A combination of linkage mapping in two F(2) populations and physical mapping with emerging genome sequence data was used to position 434 PCR-based markers including SNPs. Three-hundred-and-forty markers were used to genotype 102 tomato lines representing wild species, landraces, vintage cultivars, and contemporary (fresh market and processing) varieties. Principal component analysis confirmed genetic divergence between market classes of cultivated tomato (P <0.0001). A genome-wide survey indicated that linkage disequilibrium (LD) decays over 6-8 cM when all cultivated tomatoes, including vintage and contemporary, were considered together. Within contemporary processing varieties, LD decayed over 6-14 cM, and decay was over 3-16 cM within fresh market varieties. Significant inter-chromosomal (gametic phase) LD was detected in both fresh market and processing varieties between chromosomes 2 and 3, and 2 and 4, but in distinct chromosomal locations for each market class. Additional LD was detected between chromosomes 3 and 4, 3 and 11, and 4 and 6 in fresh market varieties and chromosomes 3 and 12 in processing varieties. These results suggest that breeding practices for market specialization in tomato have led to a genetic divergence between fresh market and processing types.

Identification of Soybean Genotypes Resistant to Cercospora sojina by Field Screening and Molecular Markers.

Frogeye leaf spot (FLS) of soybean, caused by Cercospora sojina, has been a problem in the southern United States for many years and has recently become a greater problem in the northern United States. Cultivars resistant to FLS have been developed for planting in the southern United States and resistance in many of these cultivars is conditioned by the Rcs3 gene. This gene conditions immunity to all known races and isolates of the pathogen. Resistance to C. sojina in soybean genotypes (cultivars and breeding lines) adapted to north-central U.S. production region is unknown. The objectives of this study were to (i) identify maturity group (MG) III, IV, and V soybean genotypes resistant to C. sojina race 11 by field screening at multiple locations over years and (ii) determine whether FLS resistance in these genotypes is likely to be conditioned by the Rcs3 gene. In total, 1,350 genotypes were evaluated for resistance to race 11 in field trials, and 13 MG III, 45 MG IV, and 15 MG V genotypes did not develop symptoms of FLS. Of these, 54 were subsequently tested for the possible presence of Rcs3 using five molecular markers located within 2 centimorgans (cM) of the gene. None of the MG III genotypes tested had the Rcs3 haplotype of cv. Davis, the source of Rcs3; six of the MG IV genotypes and seven of the MG V genotypes had the Rcs3 haplotype. This is the first report of the presence of the Rcs3 haplotype in LN 97-15076 and S99-2281. The soybean genotypes predicted to have the Rcs3 gene and other genotypes with no FLS symptoms in field trials may be useful in developing soybean cultivars with broad resistance to FLS and adapted to the northern United States.

The Fusarium wilt resistance locus Fom-2 of melon contains a single resistance gene with complex features.

The soil-borne fungus Fusarium oxysporum f.sp. melonis causes significant losses in the cultivated melon, a key member of the economically important family, the Cucurbitaceae. Here, we report the map-based cloning and characterization of the resistance gene Fom-2 that confers resistance to race 0 and 1 of this plant pathogen. Two recombination events, 75 kb apart, were found to bracket Fom-2 after screening approximately 1324 gametes with PCR-based markers. Sequence analysis of the Fom-2 interval revealed the presence of two candidate genes. One candidate gene showed significant similarity to previously characterized resistance genes. Sequence analysis of this gene revealed clear polymorphisms between resistant and susceptible materials and was therefore designated as Fom-2. Analysis of susceptible breeding lines (BL) presenting a haplotype very similar to the resistant cultivar MR-1 indicated that a gene conversion had occurred in Fom-2, resulting in a significant rearrangement of this gene. The second candidate gene which shared high similarity to an essential gene in Arabidopsis, presented an almost identical sequence in MR-1 and BL, further supporting Fom-2 identity. The gene conversion in Fom-2 produced a truncated R gene, revealing new insights into R gene evolution. Fom-2 was predicted to encode an NBS-LRR type R protein of the non-TIR subfamily. In contrast to most members of this class a coiled-coil structure was predicted within the LRR region rather than in the N-terminal. The Fom-2 physical region contained retroelement-like sequences and truncated genes, suggesting that this locus is complex.

Comparative mapping and marker-assisted selection in Rosaceae fruit crops.

The development of saturated linkage maps using transferable markers, restriction fragment length polymorphisms, and micro-satellites has provided a foundation for fruit tree genetics and breeding. A Prunus reference map with 562 such markers is available, and a further set of 13 maps constructed with a subset of these markers has allowed genome comparison among seven Prunus diploid (x = 8) species (almond, peach, apricot, cherry, Prunus ferganensis, Prunus davidiana, and Prunus cerasifera); marker colinearity was the rule with all of them. Preliminary results of the comparison between apple and Prunus maps suggest a high level of synteny between these two genera. Conserved genomic regions have also been detected between Prunus and Arabidopsis. By using the data from different linkage maps anchored with the reference Prunus map, it has been possible to establish, in a general map, the position of 28 major genes affecting agronomic characters found in different species. Markers tightly linked to the major genes responsible for the expression of important traits (disease/pest resistances, fruit/nut quality, self-incompatibility, etc.) have been developed in apple and Prunus and are currently in use for marker-assisted selection in breeding programs. Quantitative character dissection using linkage maps and candidate gene approaches has already started. Genomic tools such as the Prunus physical map, large EST collections in both Prunus and Malus, and the establishment of the map position of high numbers of ESTs are required for a better understanding of the Rosaceae genome and to foster additional research and applications on fruit tree genetics.