The inheritance of resistance to bacterial leaf spot of lettuce caused by Xanthomonas campestris pv. vitians in three lettuce cultivars.

Lettuce yields can be reduced by the disease bacterial leaf spot (BLS) caused by the pathogen Xanthomonas campestris pv. vitians (Xcv) and host resistance is the most feasible method to reduce disease losses. The cultivars La Brillante, Pavane and Little Gem express an incompatible host-pathogen interaction as a hypersensitive response (HR) to California strains of Xcv resulting in resistance. Little was known about the inheritance of resistance; however, resistance to other lettuce pathogens is often determined by resistance gene candidates (RGCs) encoding nucleotide-binding leucine-rich repeat (NB-LRR) proteins. Therefore, we determined the inheritance of BLS resistance in the cultivars La Brillante, Little Gem and Pavane and mapped it relative to RGCs. The reaction to Xcv was analyzed in nine F1, F2 and recombinant inbred line populations of lettuce from HR×compatible or HR×HR crosses. The HR in La Brillante, Pavane and Little Gem is conditioned by single dominant genes, which are either allelic or closely linked genes. The resistance gene in La Brillante was designated Xanthomonas resistance 1 (Xar1) and mapped to lettuce linkage group 2. Xar1 is present in a genomic region that contains numerous NB-LRR encoding RGCs and functional pathogen resistance loci in the RGC2 family. The Xar1 gene confers a high level of BLS resistance in the greenhouse and field that can be introgressed into commercial lettuce cultivars to reduce BLS losses using molecular markers

Development and application of a 6.5 million feature Affymetrix Genechip® for massively parallel discovery of single position polymorphisms in lettuce (Lactuca spp.)

BACKGROUND: High-resolution genetic maps are needed in many crops to help characterize the genetic diversity that determines agriculturally important traits. Hybridization to microarrays to detect single feature polymorphisms is a powerful technique for marker discovery and genotyping because of its highly parallel nature. However, microarrays designed for gene expression analysis rarely provide sufficient gene coverage for optimal detection of nucleotide polymorphisms, which limits utility in species with low rates of polymorphism such as lettuce (Lactuca sativa). RESULTS: We developed a 6.5 million feature Affymetrix GeneChip® for efficient polymorphism discovery and genotyping, as well as for analysis of gene expression in lettuce. Probes on the microarray were designed from 26,809 unigenes from cultivated lettuce and an additional 8,819 unigenes from four related species (L. serriola, L. saligna, L. virosa and L. perennis). Where possible, probes were tiled with a 2 bp stagger, alternating on each DNA strand; providing an average of 187 probes covering approximately 600 bp for each of over 35,000 unigenes; resulting in up to 13 fold redundancy in coverage per nucleotide. We developed protocols for hybridization of genomic DNA to the GeneChip® and refined custom algorithms that utilized coverage from multiple, high quality probes to detect single position polymorphisms in 2 bp sliding windows across each unigene. This allowed us to detect greater than 18,000 polymorphisms between the parental lines of our core mapping population, as well as numerous polymorphisms between cultivated lettuce and wild species in the lettuce genepool. Using marker data from our diversity panel comprised of 52 accessions from the five species listed above, we were able to separate accessions by species using both phylogenetic and principal component analyses. Additionally, we estimated the diversity between different types of cultivated lettuce and distinguished morphological types. CONCLUSION: By hybridizing genomic DNA to a custom oligonucleotide array designed for maximum gene coverage, we were able to identify polymorphisms using two approaches for pair-wise comparisons, as well as a highly parallel method that compared all 52 genotypes simultaneously

Oxidative discolouration in whole-head and cut lettuce: biochemical and environmental influences on a complex phenotype and potential breeding strategies to improve shelf-life

Lettuce discolouration is a key post-harvest trait. The major enzyme controlling oxidative discolouration has long been considered to be polyphenol oxidase (PPO) however, levels of PPO and subsequent development of discolouration symptoms have not always correlated. The predominance of a latent state of the enzyme in plant tissues combined with substrate activation and contemporaneous suicide inactivation mechanisms are considered as potential explanations for this phenomenon. Leaf tissue physical properties have been associated with subsequent discolouration and these may be influenced by variation in nutrient availability, especially excess nitrogen and head maturity at harvest. Mild calcium and irrigation stress has also been associated with a reduction in subsequent discolouration, although excess irrigation has been linked to increased discolouration potentially through leaf physical properties. These environmental factors, including high temperature and UV light intensities, often have impacts on levels of phenolic compounds linking the environmental responses to the biochemistry of the PPO pathway. Breeding strategies targeting the PALand PPOpathway biochemistry and environmental response genes are discussed as a more cost-effective method of mitigating oxidative discolouration then either modified atmosphere packaging or post-harvest treatments, although current understanding of the biochemistry means that such programs are likely to be limited in nature and it is likely that they will need to be deployed alongside other methods for the foreseeable future

Identification and mapping of new genes for resistance to downy mildew in lettuce

Key messageEleven new major resistance genes for lettuce downy mildew were introgressed from wild Lactuca species and mapped to small regions in the lettuce genome. Downy mildew, caused by the oomycete pathogen Bremia lactucae Regel, is the most important disease of lettuce (Lactuca sativa L.). The most effective method to control this disease is by using resistant cultivars expressing dominant resistance genes (Dm genes). In order to counter changes in pathogen virulence, multiple resistance genes have been introgressed from wild species by repeated backcrosses to cultivated lettuce, resulting in numerous near-isogenic lines (NILs) only differing for small chromosome regions that are associated with resistance. Low-pass, whole genome sequencing of 11 NILs was used to identify the chromosome segments introgressed from the wild donor species. This located the candidate chromosomal positions for resistance genes as well as additional segments. F2 segregating populations derived from these NILs were used to genetically map the resistance genes to one or two loci in the lettuce reference genome. Precise knowledge of the location of new Dm genes provides the foundation for marker-assisted selection to breed cultivars with multiple genes for resistance to downy mildew

High resolution genetic dissection of the major QTL for tipburn resistance in lettuce, Lactuca sativa

Tipburn is an important physiological disorder of lettuce, Lactuca sativa L., related to calcium deficiency that can result in leaf necrosis and unmarketable crops. The major quantitative trait locus (QTL), qTPB5.2, can account for up to 70% of the phenotypic variance for tipburn incidence in the field. This QTL was genetically dissected to identify candidate genes for tipburn by creating lines with recombination events within the QTL and assessing their resistance to tipburn. By comparing lines with contrasting haplotypes, the genetic region was narrowed down to ∼877 Kb that was associated with a reduction of tipburn by ∼60%. Analysis of the lettuce reference genome sequence revealed 12 genes in this region, one of which is a calcium transporter with a single nucleotide polymorphism in an exon between haplotypes with contrasting phenotypes. RNA-seq analysis of recombinants revealed two genes that were differentially expressed between contrasting haplotypes consistent with the tipburn phenotype. One encodes a Teosinte branched1/Cycloidea/Proliferating Cell factor transcription factor; however, differential expression of the calcium transporter was detected. The phenotypic data indicated that there is a second region outside of the ∼877 Kb region but within the QTL, at which a haplotype from the susceptible parent decreased tipburn by 10-20%. A recombinant line was identified with beneficial haplotypes in each region from both parents that showed greater tipburn resistance than the resistant parent; this line could be used as the foundation for breeding cultivars with more resistance than is currently available

Molecular markers reliably predict post-harvest deterioration of fresh-cut lettuce in modified atmosphere packaging

Salad crops: Longer-lasting lettuce Genetic studies have shown that the rate of deterioration of cut lettuce leaves in pre-packaged salads is a highly heritable trait, governed by gene regions that could be used to breed longer-lasting varieties. Many genetic studies have aimed at breeding better varieties of lettuce (Lactuca sativa), but most have focused upon those grown for whole heads, rather than the cut leaves that are becoming increasingly popular with consumers. An international team led by Ivan Simko, of the USDA in Salinas, California, have developed a genetic assay to distinguish fast- from slow-deteriorating lettuce varieties based on a single DNA region identified in a previous study. Their marker-based test may be useful in developing lettuces that show both disease resistance during cultivation, and a longer shelf life once leaves are cut for sale

The inheritance of resistance to bacterial leaf spot of lettuce caused by Xanthomonas campestris pv. vitians in three lettuce cultivars.

Lettuce yields can be reduced by the disease bacterial leaf spot (BLS) caused by the pathogen Xanthomonas campestris pv. vitians (Xcv) and host resistance is the most feasible method to reduce disease losses. The cultivars La Brillante, Pavane and Little Gem express an incompatible host-pathogen interaction as a hypersensitive response (HR) to California strains of Xcv resulting in resistance. Little was known about the inheritance of resistance; however, resistance to other lettuce pathogens is often determined by resistance gene candidates (RGCs) encoding nucleotide-binding leucine-rich repeat (NB-LRR) proteins. Therefore, we determined the inheritance of BLS resistance in the cultivars La Brillante, Little Gem and Pavane and mapped it relative to RGCs. The reaction to Xcv was analyzed in nine F1, F2 and recombinant inbred line populations of lettuce from HR×compatible or HR×HR crosses. The HR in La Brillante, Pavane and Little Gem is conditioned by single dominant genes, which are either allelic or closely linked genes. The resistance gene in La Brillante was designated Xanthomonas resistance 1 (Xar1) and mapped to lettuce linkage group 2. Xar1 is present in a genomic region that contains numerous NB-LRR encoding RGCs and functional pathogen resistance loci in the RGC2 family. The Xar1 gene confers a high level of BLS resistance in the greenhouse and field that can be introgressed into commercial lettuce cultivars to reduce BLS losses using molecular markers

Identification of QTLs conferring resistance to downy mildew in legacy cultivars of lettuce.

Many cultivars of lettuce (Lactuca sativa L.), the most popular leafy vegetable, are susceptible to downy mildew disease caused by Bremia lactucae. Cultivars Iceberg and Grand Rapids that were released in the 18th and 19th centuries, respectively, have high levels of quantitative resistance to downy mildew. We developed a population of recombinant inbred lines (RILs) originating from a cross between these two legacy cultivars, constructed a linkage map, and identified two QTLs for resistance on linkage groups 2 (qDM2.1) and 5 (qDM5.1) that determined resistance under field conditions in California and the Netherlands. The same QTLs determined delayed sporulation at the seedling stage in laboratory experiments. Alleles conferring elevated resistance at both QTLs originate from cultivar Iceberg. An additional QTL on linkage group 9 (qDM9.1) was detected through simultaneous analysis of all experiments with mixed-model approach. Alleles for elevated resistance at this locus originate from cultivar Grand Rapids