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United States Department of Agriculture

Agricultural Research Service

Research Project: ALLIUM, CUCUMIS, AND DAUCUS GERMPLASM ENHANCEMENT, GENETICS, AND BIOCHEMISTRY
2009 Annual Report


1a.Objectives (from AD-416)
Objective 1: Determine the genetic basis of and initiate selection for carrot, onion, cucumber, and melon quality attributes influencing human nutrition and health, disease resistances, and yield and quality components, and stress tolerance in cucurbits, and perform field performance and quality trials.

Objective 2: Utilize current biotechnology to discover and evaluate genetic variation and to map agriculturally important traits in Allium, Cucurbit, and Daucus germplasm, and to develop genetic and breeding stocks.

Sub-objective 2.A. Construct genetic maps of nuclear and organellar genomes using candidate genes, SCARs, SSRs, SNPs, transposon insertions, BACs, and cytogenetic stocks.

Sub-objective 2.B. Fine map pigment and carbohydrate genes in carrot and onion, resistance genes for nematode in carrot and viruses in cucurbits, and epistasis, yield and quality components in cucumber.

Sub-objective 2.C. Perform marker-assisted selection of carrot nematode resistance, onion male sterility, and cucurbit yield and quality.

Sub-objective 2.D. Evaluate transgene escape in cucurbits.

Sub-objective 2.E. Determine transposon mobility in carrot.


1b.Approach (from AD-416)
The long-term potential for improving a crop is only as great as the breadth of diversity that breeders utilize. Objective 1 targets evaluation and genetic characterization of carrot, onion, cucumber, and melon germplasm for traits important to growers and consumers.

Discovery Goal 1 - Identify unique phenotypic variation in germplasm collections and breeding stocks to improve nutritional and processing quality, disease resistance, stress tolerance, and yield of Allium, Cucurbit, and Daucus vegetables, genetically characterize observed variation and initiate genetic incorporation of these phenotypes into elite germplasms.

Many biotechnological tools have been developed to improve the efficiency of crop improvement. Objective 2 evaluates and develops these tools of carrot, onion, cucumber, and melon improvement. Identify adequate DNA polymorphisms in elite onion, cucumber, melon, and carrot germplasm to construct genetic maps for marker-facilitated selection of major horticultural traits.

Discovery Goal 2.A – Identify adequate DNA polymorphisms in elite onion, cucumber, melon, and carrot germplasm to construct genetic maps for marker-facilitated selection of major horticultural traits.

Discovery Goal 2.B – Evaluate variation at candidate genes in pigment and carbohydrate biochemical pathways for mapping in onion, cucurbit, and carrot.

Discovery Goal 2.C – Identify and utilize markers to accurately identify desirable genotypes for male sterility restoration in onion, cucurbit yield, and carrot nematode resistance.

Discovery Goal 2.D – Appraise the potential benefit(s) that transgenes might confer on transgenic populations using the ELISA test to estimate the degree of viral infection in wild populations and to determine the potential risk of virus gene introgression from commercial transgenic cultivation.

Discovery Goal 2.E - Determine if native transposable elements in the carrot genome, such as DcMaster, and introduced ones, such as maize elements Ac and Ds transpose to new chromosomal regions.


3.Progress Report
Germplasm was collected and evaluation of phenotypic and molecular relationships was made of Allium Cucumis, and Daucus germplasm, and reported in Germplasm Resources Information Network (GRIN). Wild carrot from South America and North Africa, and garlic from Uzbekistan was found to be intercrossable with carrot, and garlic, and useful traits were identified. Field evaluation of carrot, onion, and cucumber breeding stocks and experimental hybrids developed by this program was carried out in California, Michigan, Oregon, Washington, and Wisconsin. Identification of Quantitative trail locus (QTL) for melon orange pigments were identified. Genetic markers were developed to identify diverse carrots and garlic, and the locations of a carrot transposable element were mapped. Genomic information was derived from carrot bacterial artificial chromosomes (BACs) to better characterize the carrot genome.

Onion families segregating for health-enhancing fructans demonstrated that two major chromosome regions control the accumulation of health-enhancing fructans. Low-pungency onions possess recessive alleles at these two regions, reducing the ability of these onions to accumulate fructans. Evaluation of onion families for male-fertility restoration was completed and linkage to molecular markers assessed. Development of new onion families was initiated for reduced leaf waxiness (associated with tolerance to thrips), dry matter production, and bulb colors. Work continues on the identification of gene conditioning resistance to Zucchini Yellow Mosaic Virus (ZYMV) and the genetic basis of mitochondrial sorting in cucumber. These markers will be used to more efficiently develop lines for commercial production. This will shorten time for hybrid development to reduce development costs and increase grower competitiveness.

In cucumber, several F2 populations have been developed for genetic mapping of horticulturally or biologically important genes. Cucumber genomic resources are being developed (whole genome sequencing and transcriptome sequencing). The genetic diversity among USDA collection of African horned cucumber, a relative of cucumber was evaluated with molecular markers. Also, a national wide survey was conducted to identify research priorities in the public sector.


4.Accomplishments
1. Characterization of diverse carrot germplasm. Plant germplasm (seeds from diverse wild and domesticated plants from around the world) is the source of new traits valuable to crop breeders, growers, and consumers, but not much is known about the usefulness of some samples in the U.S. collection. Newly collected seed samples of wild relatives of carrot were evaluated from seeds found in Argentina and Tunisia and it was discovered that it may be possible to intercross the Argentine wild relative, Daucus pusillus, with carrot. Results to date indicate we have made a cross between the Tunisian wild relative, Daucus sahariensis, and carrot. These wild carrot relatives have resistance to leaf diseases and stress tolerance that may be useful for carrot growers, and the ability to breed these important traits into carrot would be very valuable for growers.

2. Characterization of a carrot genome library. The genetic code, or genome, of all organisms is very large, complex, and unique, so one way to simplify analysis is to break long regions of Deoxyribonucleic acid (DNA) into shorter pieces, called Bacterial Artificial Chromosomes (BACs). The collection of BACs from one organism is a BAC library. Upon evaluating a BAC library for carrot genes important for plant growth and development, biosynthesis of orange carotenoid and anthocyanin pigment genes important for human health, and genes unique to carrot were found. This information is useful to develop molecular tools to speed up carrot breeding and to understand how genes function.

3. Genetic control of orange fruit pigment color in melons and cucumbers, and nutritional quality improvement of these crops. Orange and yellow carotenoid pigments give orange-fleshed melons their familiar color, and orange-fleshed cucumbers an unusual color. These pigments are vitamin A precursors, alpha and beta carotene, which are important for eye health and a strong immune system. The genetic control of the quantity of these pigments was not known, but this research revealed that only a few genes control this important trait in cucumber, but many genes control the trait in melons. This information will provide information and tools useful to improve the nutritional quality of both these crops.

4. Genetic control of fructan accumulation in onion. Fructans are a good source of soluble dietary fiber and are associated with lower rates of colorectal cancers. Onion populations differ for concentrations of fructans. Two major regions in the onion Deoxyribonucleic acid (DNA) were identified that control the ability of onion to accumulate health-enhancing fructans. Results demonstrated that low-pungency onions carry recessive mutations at one or both of these regions, limiting their ability to accumulate fructans. These results explain why onions with lower fructan concentrations are also low pungency.

5. Cloning and characterization of gene conditioning resistance to Zucchini Yellow Mosaic Virus (ZYMV) in watermelon. ZYMV causes major losses in watermelon. The recessively inherited gene that conditions resistance to ZYMV in watermelon was cloned and characterized. The gene is eukaryotic initiation factor 4E. Mutations in this gene cause changes in this protein that block recognition of the viral Ribonucleic acid (RNA) by the plants protein synthesis machinery. These results enable watermelon breeders to select ZYMV resistance without doing laborious virus inoculations and allow for more efficient introgression of the recessively inherited ZYMV resistance into watermelon breeding lines and populations.

6. Development of cucumber genomic resources for cucumber. Although cucumber is an important vegetable crop in the United States, available genetic and genomics resources are very limited which impede progress in cucumber breeding. Whole genome sequencing of a cucumber cultivar was performed by next-generation sequencing technology. The genome has been characterized for microsatellite sequences. Over 6,000 molecular markers have been developed. This will be an important contribution to the cucurbit research community worldwide, which will facilitate studies in genetic mapping, gene cloning, and marker-assisted plant breeding.

7. Genetic diversity in African horned cucumber Cucumber relatives like the African horned cucumber (Cucumis metuliferus) possess some useful genes which are not present in cucumber. Efficient use of these genetic resources requires a better understanding of the genetic diversity in those wild species. The genetic variation of 36 C. metuliferus accessions in the USDA collection was evaluated with molecular markers and genetic diversity of these lines was found to be relatively low. Accessions with adequate polymorphisms were identified. Information gathered from this study is useful to select appropriate accession to develop populations for genetic mapping of nematode resistance genes in this species.


6.Technology Transfer

Number of Other Technology Transfer1

Review Publications
Bartoszewski, G., Gawronski, P., Szklarczyk, M., Verbakel, H., Havey, M.J. 2009. A One-Megabase Physical Map Provides Insights on Gene Organization in the Enormous Mitochondrial Genome of Cucumber. Genome. 52:299-307.

Iovene, M., Wielgus, S.M., Simon, P.W., Buell, C.R., Jiang, J. 2008. Chromatin structure and physical mapping of chromosome 6 of potato and comparative analyses with tomato. Genetics. 180(3):1307-1317.

Mccallum, J., Thomson, S., Pither-Joyce, M., Kenel, F., Clarke, A., Havey, M.J. 2008. Genetic Diversity Analysis and Single-nucleotide Polymorphism Marker Development in Cultivated Bulb Onion Based on Expressed Sequence Tag–Simple Sequence Repeat Markers. Journal of the American Society for Horticultural Science. 133:810-818.

Ipek, M., Ipek, A., Simon, P.W. 2008. Molecular Characterization of Kastamonu Garlic: An Economically Important Garlic Clone in Turkey. Scientia Horticultureae. 115(2):203-208.

Simon, P.W., Cavagnaro, P.F., Chung, S., Szklarczyk, M., Grzebelus, D., Senalik, D.A., Atkins, A.E. 2009. Characterization of a deep-coverage carrot (Daucus carota L.) BAC library and initial analysis of BAC-end sequences. Molecular Genetics and Genomics. 281(3):273-288.

Simon, P.W., Pollak, L.M., Clevidence, B.A., Holden, J.M., Haytowitz, D.B. 2009. Plant Breeding for Human Nutritional Quality. Plant Breeding Reviews. 31:325-415.

Cuevas, H.E., Staub, J.E., Simon, P.W., McCreight, J.D., Zalapa, J.E. 2008. Mapping of Genetic Loci that Regulate Accumulation of Beta-Carotene in Fruit of U.S. Western Shipping Melon (Cucumis Melo L.) and Their Association With Putative Carotenoid Biosynthesis Genes. Theoretical and Applied Genetics 117:1345-1359.

Grzebelus, D., Simon, P.W. 2008. Diversity of DcMaster-like elements of the PIF/Harbinger superfamily in the carrot genome. Genetica. Available: http://www.ncbi.nlm.nih.gov/pubmed/18535910.

Khar, A., Jakse, J., Havey, M.J. 2008. Segregations for Onion-Bulb Colors Reveal that Red is Controlled by at Least Three Loci. Journal of the American Society for Horticultural Science. 133:42–47.

Meyer, J.D., Deleu, W., Garcia-Mas, J., Havey, M.J. 2008. Construction of a Fosmid Library of Cucumber (Cucumis sativus) and Comparative Analyses of the eIF4E and eIF(iso)4E Regions from Cucumber and Melon (Cucumis melo). Molecular Genetics and Genomics. 279(5):473-480.

Al-Faifi, S., Meyer, J.D., Garcia-Mas, J., Monforte, A.J., Havey, M.J. 2008. Exploiting Synteny in Cucumis for Mapping of Psm, A Unique Locus Controlling Paternal Mitochondrial Sorting. Theoretical and Applied Genetics. 117(4):523-529.

Last Modified: 11/23/2014
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