Iraida Amaya currently works at the Department of Genomics and Biotechnology, Instituto de Investigación y Formación Agraria y Pesquera. Iraida does research in Agricultural Plant Science, Biotechnology and Genetics. Their current project is 'Improving the stability of high-quality traits of berry in different environments and cultivation systems for the benefit of European farmers and consumers. Entidad financiadora: EU H2020 SFS-2015, GoodBerry: 679303'
Skills and Expertise
PCRGene ExpressionMolecular BiologyGenomicsGeneticsBiotechnologyPlant BiotechnologyPlant BiologyGenetic EngineeringPlant GeneticsPlant BreedingGenetic AnalysisGenotypingPlant Molecular BiologyGenetic DiversityPlantsMolecular MarkersAgricultural BiotechnologyTransgenicsPlant GenomicsPlant DNA ExtractionCrop ImprovementMolecular Plant BreedingMicrosatellitesFruit QualityMarker Assisted SelectionSSRMolecular BreedingQuantitative GeneticsPlant PhenotypingCTAB DNA extractionMolecular Marker DevelopmentQuantitative Trait LociQuantitative Trait Loci MappingLinkage AnalysisDNA FingerprintsDNA Marker TechnologyQuantitative Trait Loci AnalysisSSR AnalysisDevelopment Of Molecular MarkerseQTLStrawberry
Activities will be carried out around the following lines: - Efficacy of chemical and non-chemicals soil desinfestation techniques on strawberry soil-borne pathogens control. Effect on Trichoderma wild population in soil. Influence on yield - Resistance/tolerance of strawberry cultivars to diseases and pests. Characterization of Macrophomina phaseolina isolates (strawberry and blueberry). - Berries plant problems in Huelva: main harmful agents, sector view and prospecting in affected areas.
Strawberry (Fragaria × ananassa) fruit flavor and aroma are among the most appreciated quality traits by consumers; as such, the improvement of strawberry flavor is receiving increasing priority in breeding programs. However, breeding for better aroma, particularly in strawberry as a polyploid crop, presents many challenges and requires new knowledge of the genetic determinants controlling its variation. Dr. Amaya and coworkers have identified 70 QTLs controlling the content of 48 different volatiles, including the majority of those described as key for strawberry aroma. In IOF FLAVOR we will develop a system biology approach in order to investigate the molecular mechanisms underlying some of the major and more significant QTLs. An integrated and multidisciplinary program will involve a 2 years visit of Dr Iraida Amaya to the University of Florida. She will be trained there in complementary and advanced technologies, such as saturated genetics maps of strawberry based in SNPs, global transcript quantification by RNAseq and parallel QRT-PCR, metabolomics, biochemistry and molecular biology tools, including the development of transgenic strawberry plants for validation of the identified genes. Data obtained from the different approaches will be put together and integrated to unravel the complex network of aroma compound biosynthesis. This proposal will identify genes controlling important volatiles and validate whether markers linked to them are useful in increasing the efficiency and precision of strawberry breeding programs and in the development of new cultivars with enhanced flavor. Hence a dissemination of results is planned not only to the return host’s breeding program but also towards breeding companies and the general public. The project will contribute to scientific capabilities within ERA for study of the genetic basis of fruit aroma, and will substantially enhance the career potential of Dr Amaya with state-of-the-art scientific and complementary skills.
Research Items (76)
- Jan 2018
Food fortification through the increase and/or modulation of bioactive compounds has become a major goal for preventing several diseases, including cancer. Here, strawberry lines of cv. Calypso transformed with a construct containing an anthocyanidin synthase (ANS) gene were produced to study the effects on anthocyanin biosynthesis, metabolism and transcriptome. Three strawberry ANS transgenic lines (ANS L5, ANS L15 and ANS L18) were analysed for phytochemical composition and total antioxidant capacity (TAC), and their fruit extracts assessed for cytotoxic effects on hepatocellular carcinoma. ANS L18 fruits had the highest levels of total phenolics and flavonoids, while those of ANS L15 had the highest anthocyanin concentration; TAC positively correlated with total polyphenol content. Fruit transcriptome was also specifically affected in the polyphenol biosynthesis and in other related metabolic pathways. Fruit extracts of all lines exerted cytotoxic effects in a dose/time-dependent manner, increasing cellular apoptosis and free radical levels, and impairing mitochondrial functionality.
RNA-seq has been used to perform global expression analysis of the achene and the receptacle at four stages of fruit ripening, and of the roots and leaves of strawberry (Fragaria × ananassa). About 967 million reads and 191 Gb of sequence were produced, using Illumina sequencing. Mapping the reads in the related genome of the wild diploid Fragaria vesca revealed differences between the achene and receptacle development program, and reinforced the role played by ethylene in the ripening receptacle. For the strawberry transcriptome assembly, a de novo strategy was followed, generating separate assemblies for each of the ten tissues and stages sampled. The Trinity program was used for these assemblies, resulting in over 1.4 M isoforms. Filtering by a threshold of 0.3 FPKM, and doing Blastx (E-value < 1 e-30) against the UniProt database of plants reduced the number to 472,476 isoforms. Their assembly with the MIRA program (90% homology) resulted in 26,087 contigs. From these, 91.34 percent showed high homology to Fragaria vesca genes and 87.30 percent Fragaria iinumae (BlastN E-value < 1 e-100). Mapping back the reads on the MIRA contigs identified polymorphisms at nucleotide level, using FREEBAYES, as well as estimate their relative abundance in each sample.
Flavor improvement is currently one of the most important goals for strawberry breeders. At the same time, it is one of the most complex traits to improve, involving the balanced combination of several desired characteristics such as high sweetness, moderate acidity, and the appropriate combination of aroma compounds that are beginning to be delineated in consumer tests. DNA-informed breeding will expedite the selection of complex traits, such as flavor, over traditional phenotypic evaluation, particularly when markers linked to several traits of interests are combined during the breeding process. Natural variation in mesifurane and γ-decalactone, two key volatile compounds providing sweet Sherry and fresh peach-like notes to strawberry fruits, is controlled by the FaOMT and FaFAD1 genes, respectively. In this study, we have optimized a simple PCR test for combined analysis of these genes and determined a prediction accuracy above 91% using a set of 71 diverse strawberry accessions. This high accuracy in predicting the presence of these important volatiles combined with the simplicity of the analytical methodology makes this DNA test an efficient tool for its implementation in current strawberry-breeding programs for the selection of new strawberry cultivars with superior flavor. Electronic supplementary material The online version of this article (10.1007/s11032-017-0732-7) contains supplementary material, which is available to authorized users.
Cultivated strawberry (Fragaria × ananassa) is a genetically complex allo-octoploid crop with 28 pairs of chromosomes (2n = 8x = 56) for which a genome sequence is not yet available. The diploid Fragaria vesca is considered the donor species of one of the octoploid sub-genomes and its available genome sequence can be used as a reference for genomic studies. A wide number of strawberry cultivars are stored in ex situ germplasm collections world-wide but a number of previous studies have addressed the genetic diversity present within a limited number of these collections. Here, we report the development and application of two platforms based on the implementation of Diversity Array Technology (DArT) markers for high-throughput genotyping in strawberry. The first DArT microarray was used to evaluate the genetic diversity of 62 strawberry cultivars that represent a wide range of variation based on phenotype, geographical and temporal origin and pedigrees. A total of 603 DArT markers were used to evaluate the diversity and structure of the population and their cluster analyses revealed that these markers were highly efficient in classifying the accessions in groups based on historical, geographical and pedigree-based cues. The second DArTseq platform took benefit of the complexity reduction method optimized for strawberry and the development of next generation sequencing technologies. The strawberry DArTseq was used to generate a total of 9,386 SNP markers in the previously developed ‘232’ × ‘1392’ mapping population, of which, 4,242 high quality markers were further selected to saturate this map after several filtering steps. The high-throughput platforms here developed for genotyping strawberry will facilitate genome-wide characterizations of large accessions sets and complement other available options.
Table A, List of markers mapped in the ‘232’ × ‘1392’ population, detailing adjacent sequence for DArTseq SNPs, quality scores, position in the octoploid map and in the diploid genome assembly of Tennessen et al. (2014), and genotypes in each progeny. Table B, Distribution of mapped markers in the ‘232’ × ‘1392’ map. (XLSX)
Comparison between the ‘232’ × ‘1392’ octoploid linkage map (in red) and the diploid physical map based in the genome assembly of Tennessen et al., 2014 (in green). Rearrangements are highlighted. (PDF)
Identification of the genes controlling the variation of key traits remains a challenge for plant researchers and represents a goal for the development of functional markers and their implementation in marker-assisted crop breeding. As an example we describe the identification of volatile organic compounds (VOCs) that segregate as single locus or mayor quantitative trait loci (QTL) in strawberry F1 segregating populations. Next, we describe a fast and efficient method for RNA extraction in strawberry that yields high-quality RNA for downstream RNA-seq analysis. Finally, two alternative methods for analysis of global transcript expression in contrasting lines will be described in order to identify the candidate gene and genes with differential expression using RNA-seq.
- May 2015
The receptacle of the strawberry (Fragaria × ananassa) fruit accounts for the main properties of the ripe fruit for human consumption. As it ripens, it undergoes changes similar to other fruits in sugar : acid ratio, volatile production and cell wall softening. However, the main regulators of this process have not yet been reported. The white stage marks the initiation of the ripening process, and we had previously reported a peak of expression for a FaGAMYB gene. Transient silencing of FaGAMYB using RNAi and further determination of changes in global gene expression by RNAseq, and composition of primary and secondary metabolites have been used to investigate the role played by this gene during the development of the receptacle. Down-regulation of FaGAMYB caused an arrest in the ripening of the receptacle and inhibited colour formation. Consistent with this, several transcription factors associated with the regulation of flavonoid biosynthetic pathway showed altered expression. FaGAMYB silencing also caused a reduction of ABA biosynthesis and sucrose content. Interestingly, exogenous ABA application to the RNAI-transformed receptacle reversed most defects caused by FaGAMYB down-regulation. The study assigns a key regulatory role to FaGAMYB in the initiation of strawberry receptacle ripening and acting upstream of the known regulator ABA. © 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.
Question - Does the q-PCR efficiency depend on the cDNA samples?
I think that as far as the HKG and the gene of interest have the same efficiency in each sample you don´t need to worry. You are using the HKG to normalize for differences between the different samples. However, it is important that your RNA has good 260/280 and 260/230 ratios
Background The development of a high throughput genome scanning platform for the cultivated strawberry, Fragaria ×ananassa, is necessary for enabling marker assisted breeding in this allo-octoploid species. Short-read sequencing of nineteen octoploid strawberry accessions was used to identify single nucleotide polymorphism (SNPs) and indels after alignment to the diploid Fragaria vesca ‘Hawaii 4’ reference genome and develop a 90 K Affymetrix® Axiom® array. We report the development and preliminary evaluation of this array. Results A total of ~ 36 M variants were identified in the Variant Call Format (VCF) files generated for nineteen strawberry accessions making up the Global Discovery Panel (GDP). Strategies and filtering pipelines were developed to identify markers in several different categories. Markers placed on the resulting 90 K array consisted of di-allelic SNPs (66.6%), multi-allelic SNPs (1.8%), indels (10.1%), and three categories of ploidy-reducing “haploSNPs” (11.7%), in addition to SNPs from the diploid progenitor species F. iinumae (3.9%) and speculative codon-based SNPs (5.9%). In the genotyping of 306 accessions of the cultivated octoploid strawberry, SNPs were assigned to six genotype classes with Affymetrix’ SNPolisher. The three classes of highest quality, PolyHigh Resolution (PHR), No Minor Homozygote (NMH) and Off-Target Variant (OTV) contained 25, 38 and 1% of the markers tiled on the array, respectively. These markers are highly suitable for genetic studies, as demonstrated by their use for the generation of a genetic linkage map of the full-sib family ‘Holiday’ x ‘Korona’. This map now contains 6,759 PHR SNP. NMH and OTV SNP integrate well into this PHR framework map, thus documenting the usefulness of these two classes in genotyping. Conclusions The Affymetrix® IStraw90 Axiom® array is the first high throughput genotyping platform for the allo-octoploid cultivated strawberry, and has been made commercially available to the worldwide scientific community. We expect this array to be useful for enabling marker assisted breeding, generation of high density linkage maps, identification of many QTLs for economically important traits, and genomic wide association and selection studies.
- Mar 2015
Increasing L-ascorbic acid (AsA, vitamin C) content in fruits is a common goal in current breeding programs due to its beneficial effect on human health. Attempts to increase AsA content by genetic engineering have resulted in variable success likely due to AsA's complex regulation. Here, we report the effect of ectopically expressing in tomato the D-galacturonate reductase (FaGalUR) gene from strawberry, involved in AsA biosynthesis, either under the control of the constitutive 35S or the tomato fruit-specific polygalucturonase (PG) promoters. Although transgenic lines showed a moderate increase on AsA content, complex changes in metabolites were found in transgenic fruits. Metabolomic analyses of ripe fruits identified a decrease in citrate, glutamate, asparagine, glucose and fructose, accompanied by an increase of sucrose, galactinol and chlorogenic acid. Significant metabolic changes also occurred in leaves of 35S-FaGalUR lines, which showed higher non-photochemical fluorescence quenching (NPQ), indicative of a higher constitutive photo-protective capacity. Overall, overexpression of FaGalUR increased total antioxidant capacity in fruits and the results suggest a tight control of AsA content, probably linked to a complex regulation of cellular redox state and metabolic adjustment.
- Jan 2015
Rose is one of the most economically important ornamental crops worldwide. Rosa sp. can become a model for woody ornamentals. Its genome size is relatively small (560 Mb), its genetic history with ploïdy events is well documented, and rose has a short life for a woody plant. Furthermore, different tools are available, including transcriptomic tools, genetic maps and genetic transformation protocols. Rose represents an original model for studying some ornamental traits that cannot be addressed in other model plant species such as Arabidopsis. Some of these traits, such recurrent blooming, flower morphogenesis or scent production and emission, are of economic interest. Different groups involved in rose genetics and genomics gathered to form the 'Rose Genome Sequence Initiative'. Our objective is to obtain a high quality rose genome sequence of the diploid R. chinensis 'Old Blush'. One important issue is the high level of heterozygosity of roses. To tackle this issue, different strategies are proposed: production of a haploid and development a high density genetic map to anchor the genome. This genetic map will be developed from a cross between 'Old Blush' and R. wichurana. The genotype R. chinensis 'Old Blush' will be sequenced using NGS technologies. The data will be assembled and arranged using the high-density map. In order to increase ESTs and to facilitate genome annotation, we have recently produced ESTs from various tissues of 'Old Blush' under different conditions. Digital expression (RNA Seq) was obtained from the different tissues and data are available on the following web site (https://iant.toulouse.inra.fr/plants/rosa/FATAL/). The rose genome sequence will be a great step to help identifying the molecular basis of ornamental traits and also to study genetic diversity and genome evolution in the genus Rosa and in the Rosaceae family. © 2015, International Society for Horticultural Science. All rights reserved.
Strawberry is one of the five fruit crops included in the USDA-funded multiinstitutional and trans-disciplinary project, "RosBREED: Enabling Marker- Assisted Breeding in Rosaceae". A Crop Reference Set (CRS) was developed of 900 genotypes and seedlings from 40 crosses representing the breadth of relevant diversity and encompassing founders used in breeding the domesticated strawberry. Individual native species and cultivar genotypes were included along with 10 progeny from 36 of the crosses of genotypes representing eastern and western North American and European short day and remontant cultivars. This CRS has been phenotyped in five U.S. states. Over 14 fruit quality traits have been studied, as well as remontancy, truss size, peduncle length, crop estimate, plant architecture, and disease resistance. The phenotyping conducted in the first growing season showed considerable variability amongst the genotypes and the locations for all of the characteristics. General and specific combining ability variance components were determined from the populations in order to provide breeders with guidance on the most effective breeding strategies for incorporating the superior traits from this germplasm into their programs.
Strawberry fruit flavor is the result of a complex mix of sugars, acids, and volatile organic compounds (VOCs) synthesized from a wide range of precursors including amino acids, fenylpropanoids and fatty acids. Improvement of strawberry flavor while maintaining other important traits, such as agronomic and postharvest life, requires understanding the complexity of volatile biosynthesis. We have identified genomic regions controlling the content of several VOCs using the 232 x 1392 segregating population. In order to identify the genes responsible for the variation in some of these VOCs, we are undertaking a RNA-seq approach in selected lines of the 232 x 1392 population and by sorting RNA-seq data from a second population, Mara x Elyana, based on the content of the selected VOCs. For a cluster of four phenyl-derived esters and g-dodecalactone, in LG I-1 and LGVII-1, we have identified 3 candidate genes and are using qRT-PCR, eQTL analysis and transient RNAi-induced silencing in fruits to validate their involvement. The largest cluster of QTLs controlling 16 different volatiles was mapped to LG VI-1. We have selected two contrasting bulked pools of fruits from 10 lines each with high and low levels of the six esters with QTL with stable effects higher than 30%. To identify the determinants of these variations we aim to identify differentially expressed genes between the pools using illumina RNA-seq. Preliminary results will be presented.
The RosBREED project is focused on enabling marker-assisted breeding via discovery and development of DNA tests for the most valuable QTLs that are associated with traits that are important to breeders, producers, and consumers of rosaceous crops, including strawberry. The production season for strawberries is a very narrow window of a few weeks in the Pacific Northwest and Midwest, largely dominated by short-day strawberry cultivars. While the development of remontant (day-neutral) cultivars has greatly expanded the cropping season, their cultivation in the US has thus far been limited to California, mostly due to their sensitivity to spring and summer heat. Marker-assisted selection (MAS) will facilitate the development of new remontant cultivars. The objective of this study was to validate reported associations between remontancy and existing simple sequence repeat (SSR) markers identified from the ‘Capitola’ × CF1116 (C×C) and ‘Tribute’ × ‘Honeoye’ (T×H) populations. Remontancy phenotypic data was collected in MI and OR from 2011 to 2013 for 947 cultivated strawberry accessions composed of cultivars and selections representing diverse strawberry breeding germplasm and 42 pedigreed seedling populations. Plants were considered remontant if they flowered in the Spring and after July 16th. Remontancy was also determined quantitatively as the number of weeks of flowering after July 16th. For 2013, the number of flower trusses and the number of flowers per truss were counted, cut and counted again after four weeks. The number of runners was also recorded for each accession. Genotypic data was obtained in the 947 strawberry individuals for two SSRs flanking the FaPFRU gene reported to control remontancy in C×C and from six SSRs associated with six T×H remontancy QTLs. Many strongly remontant types were discovered that could provide novel sources of heat tolerance. The microsatellite allele dosage and configuration establishment (MADCE) method, where possible, is being used to quantitatively establish the full allelic configuration at the eight SSRs. Association between marker alleles and haplotypes for these SSR markers and remontancy will be presented.
- Jul 2014
- 2014 ASHS Annual Conference
In Rosaceous crops including strawberry, the RosBREED project is focused on enabling marker-assisted breeding via identification as well as validation of markers that are associated with fruit quality traits. Around the world, strawberry is valued for its organoleptic characteristics and its nutritional content. Fruit quality traits have been prioritized in this crop. Flavor is the phenotypic result of a well synchronized combination of aroma compounds, sugars, acids and texture. The best strawberry flavor is obtained if high acids and sugars are in balance. Soluble solids content (SSC) includes mostly the concentration of sugars, followed by organic acids and soluble pectins and is positively correlated with sweetness perception and consumer likeness. The objectives of this study were: 1. to validate a previously identified QTL for SSC in 890 field-grown strawberry individuals for which SSC data were previously collected in 2011 and 2012, and 2. to identify genotypes with high SSC based on phenotypic and genotypic data. Fruit samples were collected and the resulting puree was used to measure the percent of soluble solids on a refractometer. Allele composition at the EMFv006 single sequence repeat (SSR) locus previously linked to high SSC content in the ‘Capitola’ × CF1116 strawberry population was determined after separation by capillary electrophoresis. Percent soluble solids content ranged from 3.1 to 18.7 in 2011 and between 3.7 and 16.6 in 2012 in the Oregon, and Michigan field-grown individuals evaluated. Average SSC observed in MI and OR was 11% and 8.1%, respectively. The microsatellite allele dosage and configuration establishment (MADCE) method is being used to quantitatively establish the full allelic configuration at EMFv006. We will report association of the marker alleles and haplotypes with SSC content in strawberry.
Understanding the basis for volatile organic compound (VOC) biosynthesis and regulation is of great importance for the genetic improvement of fruit flavor. Lactones constitute an essential group of fatty acid-derived VOCs conferring peach-like aroma to a number of fruits including peach, plum, pineapple and strawberry. Early studies on lactone biosynthesis suggest that several enzymatic pathways could be responsible for the diversity of lactones, but detailed information on them remained elusive. In this study, we have integrated genetic mapping and genome-wide transcriptome analysis to investigate the molecular basis of natural variation in γ-decalactone content in strawberry fruit. As a result, the fatty acid desaturase FaFAD1 was identified as the gene underlying the locus at LGIII-2 that controls γ-decalactone production in ripening fruit. The FaFAD1 gene is specifically expressed in ripe fruits and its expression fully correlates with the presence of γ-decalactone in all 95 individuals of the mapping population. In addition, we show that the level of expression of FaFAH1, with similarity to cytochrome p450 hydroxylases, significantly correlates with the content of γ-decalactone in the mapping population. The analysis of expression quantitative trait loci (eQTL) suggests that the product of this gene also has a regulatory role in the biosynthetic pathway of lactones. Altogether, this study provides mechanistic information of how the production of γ-decalactone is naturally controlled in strawberry, and proposes enzymatic activities necessary for the formation of this VOC in plants.
In an effort to implement marker-assisted breeding in Rosaceae, many traits need to be characterized in diverse germplasm. The USDA-NIFA Specialty Crop Research Initiative-funded RosBREED project includes breeding programs of four Rosaceae crops (apple, peach, cherry and strawberry). Phenotyping each crop for specific horticultural and commercial traits is an important process needed to translate genomic knowledge through marker-assisted breeding into enhanced breeding efficiency. These data will directly aid in the identification of quantitative trait loci or marker-trait associations that will be used to assist breeding programs in the future. Large-scale, standardized phenotyping protocols have been set up for each crop. The standardized phenotyping protocol for strawberries was agreed upon by the breeding teams in Oregon, Michigan, New Hampshire, California and Florida and includes four trait categories: phenology and other flower-related traits, plant characteristics, fruit characteristics, and fruit chemistry traits. We describe how each of the traits in the categories was evaluated. A summary of mean values for 37 traits of the genotypes planted at the RosBREED locations in 2011 and 2012 is provided. The phenotypic data for widely used founder germplasm that has contributed to current cultivars is available through the “Breeders Toolbox” at the Genome Database for Rosaceae (http://www.rosaceae.org/breeders_toolbox).
The RosBREED consortium has focused on developing high-throughput genome scans for apple, cherry, peach, and strawberry to facilitate quantitative trait locus (QTL) discovery and further develop the infrastructure for enabling and implementing marker-assisted breeding. This goal was met by the public release of three Illumina® Infinium® arrays for apple, peach, and cherry (8K, 9K, and 6K, respectively) that are being used by the worldwide scientific community. Development of a high-throughput genome-scanning array in strawberry has lagged behind mostly due to the challenges caused by its allo-octoploid genome. We describe the development of the International Strawberry 90K (IStraw90) Affymetrix Axiom® genotyping array. Approaches to address the allo-octoploidy challenges included a large number of SNPs (> 90,000) to compensate for potential low conversion of candidate to functional SNPs, and bioinformatic extraction of regions of reduced ploidy. This reduction of effective ploidy levels was the direct result of targeting subgenome-specific sites. We also report results of the preliminary evaluation of this array in 384 strawberry samples consisting of mapping populations, breeding populations and their founders, in addition to few diverse strawberry individuals. We expect this array to enable genome-wide scanning in the octoploid strawberry and to facilitate QTL discovery for many traits of economic significance in this important fruit crop.
In an effort to implement marker-assisted breeding in Rosaceae, many traits need to be characterized in diverse germplasm. The USDA–NIFA Specialty Crop Research Initiative-funded RosBREED project includes breeding programs of four Rosaceae crops, apple, peach, cherry, and strawberry. Among them, strawberry is the only perennial herbaceous species. Phenotyping strawberry for specific horticultural and commercial traits is an important process needed to identify genotypic marker(s) associated with specific traits. This process is the first step in translating genomic knowledge into enhanced breeding efficiency through marker-assisted breeding. Large-scale standardized phenotyping protocols have been set up for each crop. The standardized phenotyping protocol for strawberries, as agreed upon by the breeding teams in Oregon, Michigan, New Hampshire, California, and Florida, will be presented. The protocol includes four trait categories: phenology, plant characteristics, fruit characteristics, and fruit chemistry. Phenotyping the RosBREED strawberry of 947 individuals representing the breadth of relevant diversity used in breeding the domesticated strawberry, took place in 2011 and 2012. These data will be used to identify quantitative trait loci and marker trait associations that can assist breeding programs in the future. The phenotypic data for widely used founder accessions that have contributed to current cultivars is available through the “Breeders Toolbox” at the Genome Database for Rosaceae (http://www.rosaceae.org/ breeders_toolbox).
Strawberry is a soft fruit with a short postharvest shelf-life. The loss of fruit firmness during ripening is mainly due to the disassembly of parenchyma cell walls mediated by the expression of genes encoding enzymes acting on pectins, such as pectate lyase, or hemicellulose, e.g. endo-β-1,4-glucanase. To determine if the simultaneous down-regulation of FaplC and FaEG3 genes, encoding a pectate lyase and a endo-β-1,4-glucanase, respectively, exerted an additive effect on strawberry softening, transgenic plants expressing tandem antisense sequences of both genes under the control of the constitutive promoter CaMV35S were generated. Fifteen independent transgenic lines were obtained and fruit yields and several quality parameters of transgenic ripe fruit were recorded during two consecutive years. Fruit yield was reduced in most of the lines, especially in the first evaluation period, and five out of 15 lines (33 %) did not set fruit. The expression of FaplC and FaEG3 genes was measured in ripe fruits from six selected lines showing the highest fruit yields. All selected lines showed a high level of FaplC gene silencing, ranging from 97 to 71 %; however, FaEG3 gene expression was only significantly down-regulated in two lines. Fruit colour and soluble solids contents were similar in control and transgenic ripe fruits, while fruit weight was slightly lower than control in some of the lines. In all lines, transgenic fruits were significantly firmer than control, with an increase in firmness ranging from 19 to 32 %. The reduction of fruit softening in transgenic fruits was not correlated with the suppression of FaEG3 gene expression, and lines with the highest simultaneous down-regulation of FaplC and FaEG3 showed similar fruit firmness to lines where only FaplC was suppressed. These results indicate that pectate lyase and endo-β-1,4-glucanase do not act in an additive or synergistic way during strawberry softening, and question the role of glucanases in this process.
- Sep 2012
The term "vitamin" is used to define a number of organic compounds that have to be obtained from different foods because the organism itself cannot synthesize them in the quantities needed to sustain life. Vitamin C is the common name for L-ascorbic acid. In humans, the principal role of this molecule is to scavenge reactive oxygen species, due to its antioxidant capacity, and to serve as cofactor for many enzymes. A deficiency of L-ascorbic acid is traditionally linked to human diseases such as scurvy. Plant foods are the principal source of L-ascorbic acid for humans. There is a high variability of L-ascorbic acid content in the various plant organs that are used for human consumption. This diversity is related to the specific functions played by L-ascorbic acid in the different plant tissues. The net content of L-ascorbic acid in plants is determined through a balance of the activities of different biosynthetic, recycling, and catabolic pathways. Here we review the importance of L-ascorbic acid for human health, the current knowledge on its metabolism and function in plants, and the efforts that have already been made by genetic modification to improve its content in plant organs used for human food. We provide a current and forward looking perspective of how plant science can contribute to improving the L-ascorbic acid content in crop species using gene transformation, quantitative trait loci and association mapping-based approaches.
Seventy-seven olive accessions corresponding to 25 cultivars from the Extremadura region of Spain were studied using four microsatellite or SSR markers in order to fingerprint them, and evaluate genetic similarity and relationships between local and introduced olive cultivars. The number of alleles per locus ranged from 4 to 8, with a mean of 6.25 alleles per primer pair (a total of 25 alleles). The observed heterozygosity ranged from 0.58 to 0.95, while the expected heterozygosity varied between 0.68 and 0.83. The polymorphism information content values ranged from 0.63 to 0.79. The mean polymorphism information content value of 0.70 for the SSR loci provided sufficient discriminating ability to evaluate the genetic diversity among the cultivars. The SSR data allowed unequivocal identification of all the cultivars; a combination of three SSR markers was sufficient to discriminate all 25 olive cultivars. A dendrogram was prepared, using the unweighted pair-group method with arithmetic mean clustering algorithm; it depicted the pattern of relationships between the cultivars. Most of the local cultivars grouped according to their geographic origin. No clear clustering trends were observed when the morphological traits of fruit endocarps or fruit use of cultivars were employed as analysis criteria. We conclude that there is a high level of variability among local olive cultivars from the Extremadura region at both the morphological and molecular levels; these data should be useful for identifying and distinguishing local germplasm.
Improvement of strawberry (Fragaria × ananassa) fruit flavor is an important goal in breeding programs. To investigate genetic factors controlling this complex trait, a strawberry mapping population derived from genotype '1392', selected for its superior flavor, and '232' was profiled for volatile compounds over 4 years by headspace solid phase microextraction coupled to gas chromatography and mass spectrometry. More than 300 volatile compounds were detected, of which 87 were identified by comparison of mass spectrum and retention time to those of pure standards. Parental line '1392' displayed higher volatile levels than '232', and these and many other compounds with similar levels in both parents segregated in the progeny. Cluster analysis grouped the volatiles into distinct chemically related families and revealed a complex metabolic network underlying volatile production in strawberry fruit. Quantitative trait loci (QTL) detection was carried out over 3 years based on a double pseudo-testcross strategy. Seventy QTLs covering 48 different volatiles were detected, with several of them being stable over time and mapped as major QTLs. Loci controlling γ-decalactone and mesifurane content were mapped as qualitative traits. Using a candidate gene approach we have assigned genes that are likely responsible for several of the QTLs. As a proof of concept we show that one homoeolog of the O-methyltransferase gene (FaOMT) is the locus responsible for the natural variation of mesifurane content. Sequence analysis identified 30 bp in the promoter of this FaOMT homoeolog containing putative binding sites for basic/helix-loop-helix, MYB, and BZIP transcription factors. This polymorphism fully cosegregates with both the presence of mesifurane and the high expression of FaOMT during ripening.
Breeding for fruit quality traits in strawberry (Fragaria × ananassa, 2n = 8x = 56) is complex due to the polygenic nature of these traits and the octoploid constitution of this species. In order to improve the efficiency of genotype selection, the identification of quantitative trait loci (QTL) and associated molecular markers will constitute a valuable tool for breeding programs. However, the implementation of these markers in breeding programs depends upon the complexity and stability of QTLs across different environments. In this work, the genetic control of 17 agronomical and fruit quality traits was investigated in strawberry using a F(1) population derived from an intraspecific cross between two contrasting selection lines, '232' and '1392'. QTL analyses were performed over three successive years based on the separate parental linkage maps and a pseudo-testcross strategy. The integrated strawberry genetic map consists of 338 molecular markers covering 37 linkage groups, thus exceeding the 28 chromosomes. 33 QTLs were identified for 14 of the 17 studied traits and approximately 37% of them were stable over time. For each trait, 1-5 QTLs were identified with individual effects ranging between 9.2 and 30.5% of the phenotypic variation, indicating that all analysed traits are complex and quantitatively inherited. Many QTLs controlling correlated traits were co-located in homoeology group V, indicating linkage or pleiotropic effects of loci. Candidate genes for several QTLs controlling yield, anthocyanins, firmness and L-ascorbic acid are proposed based on both their co-localization and predicted function. We also report conserved QTLs among strawberry and other Rosaceae based on their syntenic location.
Plants have several L-ascorbic acid (AsA) biosynthetic pathways, but the contribution of each one to the synthesis of AsA varyies between different species, organs, and developmental stages. Strawberry (Fragaria×ananassa) fruits are rich in AsA. The pathway that uses D-galacturonate as the initial substrate is functional in ripe fruits, but the contribution of other pathways to AsA biosynthesis has not been studied. The transcription of genes encoding biosynthetic enzymes such as D-galacturonate reductase (FaGalUR) and myo-inositol oxygenase (FaMIOX), and the AsA recycling enzyme monodehydroascorbate reductase (FaMDHAR) were positively correlated with the increase in AsA during fruit ripening. Fruit storage for 72 h in a cold room reduced the AsA content by 30%. Under an ozone atmosphere, this reduction was 15%. Ozone treatment increased the expression of the FaGalUR, FaMIOX, and L-galactose-1-phosphate phosphatase (FaGIPP) genes, and transcription of the L-galactono-1,4-lactone dehydrogenase (FaGLDH) and FAMDHAR genes was higher in the ozone-stored than in the air-stored fruits. Analysis of AsA content in a segregating population from two strawberry cultivars showed high variability, which did not correlate with the transcription of any of the genes studied. Study of GalUR protein in diverse cultivars of strawberry and different Fragaria species showed that a correlation between GalUR and AsA content was apparent in most cases, but it was not general. Three alleles were identified in strawberry, but any sequence effect on the AsA variability was eliminated by analysis of the allele-specific expression. Taken together, these results indicate that FaGalUR shares the control of AsA levels with other enzymes and regulatory elements in strawberry fruit.
Cultivated strawberry (Fragaria×ananassa) together with other economically important genera such as Rosa (roses) and Rubus (raspberry and blackberry) belongs to the subfamily Rosoideae. There is increasing interest in the development of transferable markers to allow genome comparisons within the Rosaceae family. In this report, 122 new genic microsatellite (SSR) markers have been developed from cultivated strawberry and its diploid ancestor Fragaria vesca. More than 77% of the sequences from which the markers were developed show significant homology to known or predicted proteins and more than 92% were polymorphic among strawberry cultivars, representing valuable markers in transcribed regions of the genome. Sixty-three SSRs were polymorphic in the diploid Fragaria reference population and were bin-mapped together with another five previously reported but unmapped markers. In total, 72 loci were distributed across the seven linkage groups. In addition, the transferability of 174 Fragaria SSRs to the related Rosa and Rubus genera was investigated, ranging from 28.7% for genic-SSRs in rose to 16.1% for genomic-SSRs in raspberry. Among these markers, 33 and 16 were both localized in the diploid Fragaria reference map and cross-amplified in rose and raspberry, respectively. These results indicate that transferability of SSRs across the Rosoideae subfamily is limited. However, we have identified a set of Fragaria markers, polymorphic in the diploid reference population, which cross-amplified in both Rosa and Rubus, which represents a valuable tool for comparative mapping and genetic diversity analyses within the Rosoideae subfamily. KeywordsComparative mapping–Synteny–Strawberry–Rose–Raspberry–Rosoideae
The conservation and characterization of genetic resources have increasing importance as a source of variability in breeding programmes and to preserve biodiversity. In order to establish a refreshed collection free of pathogens at the IFAPA Strawberry Germplasm Collection we have initiated the detection of pathogens during the refreshment process, a sanitary control of plant material inputs and outputs, and a specific analyses of symptomatic plants observed in the in vivo collection. To isolate and identify pathogenic fungi and oomycetes, we are applying conventional techniques. Both molecular and immune-serological diagnoses are being used for detection of nematode-transmitted viruses (ArMV, TBRV, RpRSV, SLRSV) and only molecular techniques for aphid-transmitted viruses (SVBV, SMoV, SCV and SMYEV). Results show that most of the refreshed material is free of lethal fungi, oomycetes or viruses. However, on symptomatic plants, we detected the species Phytophthora cactorum and other plant pathogenic genera such as Cylindrocarpon, Fusarium, Pestalotia and Rhizoctonia, whose pathogenicity to strawberry needs to be characterised. Exceptionally, SMoV and SMYEV viruses have been intercepted in imported material. For the collection management, sanitary measures to avoid pathogen establishment and dispersal within the collection are being adopted.
Unlike other important crops analyzed so far for genetic diversity and population structure, the brief history and particularities of the genetics of the cultivated strawberry (Fragaria ×ananassa Duchesne) have limited its genetic characterization. The genomic composition and the pattern of inheritance have not been fully elucidated, although a number of studies have suggested a highly diploidized genome. In this study, the similarity relationships and structure of 92 selected strawberry cultivars with widely diverse origins have been established using simple sequence repeat (SSR) markers derived from expressed sequence tags (EST-SSR markers). Genetic analysis performed by the unweighted pair group method with arithmetic mean clustering revealed a distribution according to both date of cultivar release and breeding for a specific climatic adaptation. Additionally, a model-based clustering approach identified three populations among the strawberry cultivars with an overall FST value of 0.15 to 0.16. Both analyses support a limited differentiation of modern cultivars, most probably as a consequence of the methodology of strawberry breeding. Interestingly, the collection of strawberry cultivars here analyzed showed comparable genetic differentiation to that observed in natural populations of Fragaria chiloensis (L.) Mill., one of its wild ancestors. Our results suggest that breeding has produced a small but significant reduction on the genetic diversity of F. ×ananassa. The panel of 10 EST-SSRs described in this work provided an extremely low probability of confusion (less than 10-11), offering an efficient and accurate method for cultivar identification.
- Jan 2009
Unlike other important crops analyzed so far for genetic diversity and population structure, the brief history and particularities of the genetics of the cultivated strawberry (Fragaria ·ananassa Duchesne) have limited its genetic characterization. The genomic composition and the pattern of inheritance have not been fully elucidated, although a number of studies have suggested a highly diploidized genome. In this study, the similarity relationships and structure of 92 selected strawberry cultivars with widely diverse origins have been established using simple sequence repeat (SSR) markers derived from expressed sequence tags (EST-SSR markers). Genetic analysis performed by the unweighted pair group method with arithmetic mean clustering revealed a distribution according to both date of cultivar release and breeding for a specific climatic adaptation. Additionally, a model-based clustering approach identified three populations among the strawberry cultivars with an overall FST value of 0.15 to 0.16. Both analyses support a limited differentiation of modern cultivars, most probably as a consequence of the methodology of strawberry breeding. Interestingly, the collection of strawberry cultivars here analyzed showed comparable genetic differentiation to that observed in natural populations of Fragaria chiloensis (L.) Mill., one of its wild ancestors. Our results suggest that breeding has produced a small but significant reduction on the genetic diversity of F. ·ananassa. The panel of 10 EST-SSRs described in this work provided an extremely low probability of confusion (less than 10–11), offering an efficient and accurate method for cultivar identification.
- Dec 2006
Microsatellite or simple sequence repeat markers derived from expressed sequence tags (ESTs) provide genetic markers within potentially functional genes, which could be very useful for breeding programs. To date, the development of microsatellite markers in the genus Fragaria has focused mainly on Fragaria vesca. However, most of the interests of breeding programs relate to specific characteristics of cultivated strawberry. Here, we describe a set of 10 EST-derived microsatellites from Fragaria × ananassa. These markers showed high levels of polymorphism within strawberry cultivars and among different Fragaria species, indicating their potential for genetic studies not only on strawberry but also in other species within the genus.
The study of mutants impaired in the sensitivity or synthesis of abscisic acid (ABA) has become a powerful tool to analyse the interactions occurring between the ABA and ethylene signalling pathways, with potential to change the traditional view of the role of ABA as just being involved in growth inhibition. The tss2 tomato mutant, which is hypersensitive to NaCl and osmotic stress, shows enhanced growth inhibition in the presence of exogenous ABA. The tos1 tomato mutant is also hypersensitive to osmotic stress, but in contrast to tss2, shows decreased sensitivity to ABA. Surprisingly, blocking ethylene signalling suppresses the growth defect of tss2 seedlings on ABA, NaCl, and osmotic stress, but not the osmotic hypersensitivity of tos1. The ethylene production of tss2 seedlings is increased compared with that of control seedlings under osmotic stress. In addition, the tss2 plants are hypersensitive to root growth inhibition by the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC). This suggests that, in addition to ABA regulation, TSS2 acts as a negative regulator of endogenous ethylene accumulation. As previously shown in Arabidopsis, it is shown here that extensive cross-talk occurs between the ABA and ethylene signalling pathways in tomato and that the TSS2 and TOS1 loci appear as regulators of this cross-talk.
The strawberry (Fragaria×ananassa) FaGAST gene encodes a small protein with 12 cysteine residues conserved in the C-terminal region similar to a group of proteins identified in other species with diverse assigned functions such as cell division, elongation, or elongation arrest. This gene is expressed in the fruit receptacle, with two peaks during ripening at the white and the red-ripe stages, both coincident with an arrest in the growth pattern. Expression is also high in the roots but confined to the cells at the end of the elongation zone. Exogenous application of gibberellin increased the transcript level of the FaGAST gene in strawberry fruits. Ectopic expression of FaGAST in transgenic Fragaria vesca under the control of the CaMV-35S promoter caused both delayed growth of the plant and fruits with reduced size. The same growth defect was observed in Arabidopsis thaliana plants overexpressing FaGAST. In addition, the transgenic plants exhibited late flowering and low sensitivity to exogenous gibberellin. Taken together, the expression pattern, the regulation by gibberellin, and the transgenic phenotypes point to a role for FaGAST in arresting cell elongation during strawberry fruit ripening.
The isolation and characterization of fruit-specific promoters are critical for the manipulation of the nutritional value and quality of fruits by genetic engineering. The analysis of regulatory sequences of many ripening-related genes has remained elusive for many species due to their low transformation efficiency and/or lengthy regeneration of a small number of transgenic plants. Strawberry is an important crop and represents one of the most widely studied non-climacteric model systems. However, until recently, its difficult regeneration has limited the functional study of promoters by stable transformation. A protocol based on biolistic transient transformation has been developed in order to study the function of promoters in a fast and efficient manner in strawberry fruits. The protocol has been applied to the study of the GalUR promoter, a gene involved in the biosynthesis of vitamin C in this fruit. The activity of the GalUR promoter is restricted to the fruit, being strictly dependent on light. The analysis of deletion series revealed the presence of a minimum activation region 397 bp upstream of the gene with a putative G-box motif, and a negative regulatory region between −397 and −518 bp, where an I-box was identified. The transient assay has been used to study the activity of the tomato polygalacturonase and the pepper fibrillin promoters in strawberry fruits. Whereas slight activity was observed with the fibrillin promoter, no significant activity was found with the polygalacturonase promoter. The GalUR promoter in transiently transformed ripe tomato fruits showed no activity, indicating the presence of regulatory sequences specific for its function in strawberry fruit.
The last step in the synthesis of lignin and suberin has been proposed to be catalyzed by peroxidases, although other proteins may also be involved. To determine which peroxidases are involved in the synthesis of lignin and suberin, five peroxidases from tomato (Lycopersicon esculentum) roots, representing the majority of the peroxidase activity in this organ, have been partially purified and characterized kinetically. The purified peroxidases with isoelectric point (pI) values of 3.6 and 9.6 showed the highest catalytic efficiency when the substrate used was syringaldazine, an analog of lignin monomer. Using a combination of transgenic expression and antibody recognition, we now show that the peroxidase pI 9.6 is probably encoded by TPX1, a tomato peroxidase gene we have previously isolated. In situ RNA hybridization revealed that TPX1 expression is restricted to cells undergoing synthesis of lignin and suberin. Salt stress has been reported to induce the synthesis of lignin and/or suberin. This stress applied to tomato caused changes in the expression pattern of TPX1 and induced the TPX1 protein. We propose that the TPX1 product is involved in the synthesis of lignin and suberin.
- Sep 1999
The cell wall is a fundamental component in the response of plants to environmental changes. To directly assess the role of the cell wall we have increased the expression and activity of a cell wall associated peroxidase (TPX2), an enzyme involved in modifying cell wall architecture. Overexpression of TPX2 had no effect on wild-type development, but greatly increased the germination rate under high salt or osmotic stress. Differential scanning calorimetry showed that transgenic seeds were able to retain more water available for germination than wild-type seeds. Thermoporometry calculations indicated that this could be due to a lower mean pore size in the walls of transgenic seeds. Therefore, the higher capacity of transgenic seeds in retaining water could result in higher germination rates in conditions where the availability of water is restricted.
Plant species exhibit two primary forms of flowering architecture, namely, indeterminate and determinate. Antirrhinum is an indeterminate species in which shoots grow indefinitely and only generate flowers from their periphery. Tobacco is a determinate species in which shoot meristems terminate by converting to a flower. We show that tobacco is responsive to the CENTRORADIALIS (CEN) gene, which is required for indeterminate growth of the shoot meristem in Antirrhinum. Tobacco plants overexpressing CEN have an extended vegetative phase, delaying the switch to flowering. Therefore, CEN defines a conserved system controlling shoot meristem identity and plant architecture in diverse species. To understand the underlying basis for differences between determinate and indeterminate architectures, we isolated CEN-like genes from tobacco (CET genes). In tobacco, the CET genes most similar to CEN are not expressed in the main shoot meristem; their expression is restricted to vegetative axillary meristems. As vegetative meristems develop into flowering shoots, CET genes are downregulated as floral meristem identity genes are upregulated. Our results suggest a general model for tobacco, Antirrhinum, and Arabidopsis, whereby the complementary expression patterns of CEN-like genes and floral meristem identity genes underlie different plant architectures.
The overall aerial architecture of flowering plants depends on a group of meristematic cells in the shoot apex. We demonstrate that the Arabidopsis TERMINAL FLOWER 1 gene has a unified effect on the rate of progression of the shoot apex through different developmental phases. In transgenic Arabidopsis plants which ectopically express TERMINAL FLOWER 1, both the vegetative and reproductive phases are greatly extended. As a consequence, these plants exhibit dramatic changes in their overall morphology, producing an enlarged vegetative rosette of leaves, followed by a highly branched inflorescence which eventually forms normal flowers. Activity of the floral meristem identity genes LEAFY and APETALA 1 is not directly inhibited by TERMINAL FLOWER 1, but their upregulation is markedly delayed compared to wild-type controls. These phenotypic and molecular effects complement those observed in the tfl1 mutant, where all phases are shortened. The results suggest that TERMINAL FLOWER 1 participates in a common mechanism underlying major shoot apical phase transitions, rather than there being unrelated mechanisms which regulate each specific transition during the life cycle.
- Jan 1996
- Physical Stresses in Plants
The plasmid pMN24 carrying a functional copy of the nitrate reductase gene Nitl was used for insertional mutagenesis of nit-Chlamydomonas reinhardtii. Over 6800 nit+ transformants were screened for mutations causing NaCl sensitivity. Four mutants with salt sensitivity that cosegregated with nit+were isolated. Salt sensitivity of these mutants also cosegregated with the presence of a DNA fragment hybridizing to the Nitl plasmid pMN24. Over 8000 EMS mutagenized yeast colonies were screened for salt sensitivity and eleven sensitive lines were isolated. One of these, SS2, was complemented for salt tolerance with a genomic library from wild type yeast. A salt tolerant transformant was found to harbor a DNA fragment encoding the calcineurin regulatory subunit (CNB1) gene. Gene disruption and segregation analysis indicated the SS2 and CNB1 loci were allelic. Molecular modification of calcineurin catalytic subunit (CNA) to remove the calmodulin (CAM)-binding and auto-inhibitory domains was performed. The subsequent truncated CNA (CNAtr1) gene, along with a CNB1 gene were over-expressed in yeast. This resulted in a 5 fold increase in salt tolerance. The CNAtr1 and CNB1 genes were also transformed into tobacco and several tobacco lines expressing yeast CNAtr1 and CNB1 transcripts were isolated. Preliminary experiments indicate that CNAtr1/CNB1 transgenic tobacco plants have altered growth on salt.
- International Plant and Animal Genome Conference XXII 2014
Understanding the basis for volatile organic compound (VOC) biosynthesis and regulation is of great importance for the genetic improvement of fruit flavor. Lactones constitute an essential group of fatty acid-derived VOCs conferring peach-like aroma to a number of fruits including peach, plum, pineapple and strawberry. Early studies on lactone biosynthesis suggest that several enzymatic pathways could be responsible for the diversity of lactones, but detailed information on them remained elusive. The content of g-decalactone is controlled by one dominant locus mapped to the bottom arm of LG III-2 in strawberry. Our laboratories unknowingly took separate yet complementary genomics approaches to simultaneously identify the fatty acid desaturase FaFAD1 as the gene underlying the locus that controls g-decalactone production in ripening fruits. The gene was discovered using genetic mapping, GC/MS and genome-wide transcriptome analysis using either in vivo or in silico bulked RNAseq expression data. The FaFAD1 gene is specifically expressed in ripe fruits and its expression fully correlates with the presence of g-decalactone in all 95 individuals of one of the mapping populations. A functional, PCR‐based molecular marker was developed that cosegregates perfectly with the phenotype in other F1 and BC1 populations. The marker also accurately predicted the volatile phenotype in germplasm outside our populations including cultivars and wild Fragaria species. Additional candidate genes of the biosynthetic pathway of lactones are also reported based on eQTL analysis. All together, these studies provide mechanistic information of how the production of g-decalactone is naturally controlled in strawberry, propose enzymatic activities necessary for the formation of this VOC in plants and deliver molecular markers that could be used for the selection of flavorful plants across the Rosaceae.
- International Plant and Animal Genome Conference XX 2012
Improvement of flavor and nutritional quality is an important objective in current strawberry breeding programs. QTLs and candidate genes have been identified for yield and related traits, and fruit quality characters evaluated during 3 years using ‘functional’ linkage maps of strawberry. 33 QTLs were identified for 14 different traits and approximately 37% of them were stable over time. 1-5 QTLs were detected per trait, with individual effects ranging from 9.2 to 30.5% of the phenotypic variation. Candidate genes for QTLs controlling different traits were identified based in their co-location and predicted function. In addition, syntenic QTLs among strawberry and other Rosaceae are proposed. Besides sugars, acids and texture, strawberry flavor is determined by a complex balance of aroma compounds. More than 300 volatile compounds were detected by HS-SPME-GC-MS in the population and 87 of them were identified. Cluster analysis grouped the volatiles into distinct chemically related families and revealed a complex metabolic network underlying volatile production in strawberry fruit. Seventy QTLs covering 48 different volatiles were detected, with several of them being stable over time and mapped as major QTLs. Using a candidate gene approach we have assigned genes that are likely responsible for several of the QTLs. As a proof of concept we show that one homoeolog of FaOMT is the locus responsible for the natural variability of mesifurane content. Sequence analysis identified a polymorphism in the promoter of this FaOMT homoeolog that fully co-segregates with both the presence of mesifurane and the high expression of FaOMT during ripening.
- International Plant and Animal Genome Conference XXI 2013
A central goal of the RosBREED and FruitBreedomics consortia has been to establish SNP arrays for peach, cherry, apple, and strawberry, to facilitate QTL discovery and marker-assisted breeding. This goal has been advanced by the release of three Illumina® Infinium® arrays for apple, peach, and cherry (8K, 9K and 6K, respectively). Here, we report on the development of a 90K Strawberry Affymetrix Axiom® and a 20K Apple Infinium® genotyping array. The cultivated strawberry is an allo-octoploid. This level and type of ploidy creates challenges which we addressed in several ways. First, the large size of the array permits success despite a lower conversion rate of candidate to functional SNPs than for diploid crops. Second, we exploited both site-specific, biological as well as designed reductions in ploidy. The array will target several polymorphism types, including indels and di- and multi-allelic SNPs. For apple, the new SNP markers are located in focal points of max 10 KB, an approach that favours the design of multi-allelic haplotypes. Focal points have an average of 8 SNPs selected to evenly represent the members of the discovery panel. Issues related to the duplicated nature of the apple genome are accounted for in several ways, including the use of double haploids. We will describe our approaches for the design of these two SNP arrays. We also report on a new bioinformatics pipeline, which includes local re-alignment around indels and polymorphism type-specific filtering strategies. Both arrays will be commercially available in early 2013.
Rose is one of the most economically important ornamental crops worldwide. Rosa sp. is well suited genus to become a model for woody ornamentals for a number of reasons: (i) its relative small genome size (500 Mb), (ii) its well documented genetic history with ploïdy events, (iii) its short life for a woody plant, and (iv) availability of different tools, including transcriptomic tools, genetic maps and genetic transformation protocols. Furthermore, the rose represents an original model for studying some ornamental traits that can not be addressed in other model species such as Arabidopsis. Some of these traits, such recurrent blooming, flower morphogenesis or scent production and emission, are of economic interest. The objective of the Rose Genome Sequence Initiative (http://rosegenome.org) is to obtain a high quality rose genome sequence of the diploid R. chinensis ‘Old Blush’. One important issue is the high heterozygosity of roses. To tackle the heterozygosity issue, we propose to develop a high density genetic map to anchor the genome. This genetic map will be developed from a cross between ‘Old Blush’ and R. wichurana. The genotype R. chinensis ‘Old Blush’ will be sequenced using NGS technologies. The data will be assembled and arranged using the high density map. New ESTs will be developed to facilitate assembly and gene annotation. The rose genome sequence will be a great step to help identifying the molecular basis of ornamental traits and also to study genetic diversity and genome evolution in the genus Rosa and in the Rosaceae family.