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ARS Home » Pacific West Area » Davis, California » Nat'l Clonal Germplasm Rep - Tree Fruit & Nut Crops & Grapes » Research » Research Project #434303

Research Project: Managing Genetic Resources and Associated Information of Grape, Tree Fruit, Tree Nut, and Other Specialty Crops Adapted to Mediterranean Climates

Location: Nat'l Clonal Germplasm Rep - Tree Fruit & Nut Crops & Grapes

2019 Annual Report

The long-term objective of this project is to preserve the genetic diversity in the collections of the 14 Mediterranean crops for current and future generations. Specifically, during the next five years we will focus on the following objectives. Objective 1: Efficiently and effectively acquire priority Mediterranean climate-adapted grape, tree fruit, tree nut, and other specialty crop genetic resources; maintain their safety, genetic integrity, health and viability; and distribute them and associated information worldwide. Objective 2: Develop more effective genetic resource maintenance, evaluation, and characterization methods and apply them to priority Mediterranean climate-adapted grape, tree fruit, tree nut, and other specialty crop genetic resources. Record and disseminate evaluation and characterization data via GRIN-Global and other data sources. Objective 3: With other NPGS genebanks and Crop Germplasm Committees, develop, update, document, and implement best management practices and Crop Vulnerability Statements for Mediterranean climate-adapted grape, tree fruit, tree nut, and other specialty crop genetic resource and information management. Basis for the objectives: The first two objectives describe the four basic tenets of the mission of the National Clonal Germplasm Repository Davis (NCGR); acquisition, maintenance, characterization and distribution of high quality, healthy, viable, true-to-type genetic resources and the associated evaluation and characterization information in GRIN-Global to the domestic and international scientific and educational communities. The third objective focuses on developing, implementing, and documenting best management practices for the collections and keeping the Crop Vulnerability Statements up-to-date. Acquisition has focused on crop wild relatives (CWRs) to fill important gaps and much of this germplasm is under quarantine and therefore in the pipeline to the Repository. Maintenance is best accomplished with young, vigorous plants and therefore the Repository is in the middle of a repropagation cycle for the collections. Distribution of germplasm to national and international researchers, breeders, and nurseries is primarily, but not limited to dormant cuttings or scionwood. Phenotypic evaluation and genetic characterization of the collections focus on understanding genetic diversity and increasing the visibility and value of the germplasm. Stakeholders are interested in phenotypic data on traits with breeding value. We use Bioversity International descriptors for germplasm evaluation and upload these data to GRIN-Global. Genetic markers discovered at the Repository and by collaborators can help clarify accession identity and assess genetic diversity, structure, and differentiation, and when combined with phenotypes will enhance the value and promote utilization of the germplasm.

Objective 1 Research Goal 1: Implement best management strategies for the acquisition, maintenance, and distribution of the genetic resources of the NCGR. Approach: This objective describes three of the four basic tenets of the mission of the National Clonal Germplasm Repository for tree fruits, nut crops and grapes (NCGR) of acquisition, maintenance of the germplasm, and distribution of high quality, healthy, viable, trueto-type genetic resources to the domestic and international scientific and educational communities. The fourth tenet is evaluation and characterization covered in Objective 2. This and other objectives are accomplished by 10 full-time staff, as well as 2-5 part-time student workers. Objective 2 Research Goal 2: To have better maintained and more thoroughly evaluated and characterized collections and to make the resulting descriptor information more complete and up-to-date in GRIN-Global. Approach: This objective describes making collection maintenance more effective and is well aligned with the maintenance portion of Objective 1. Evaluation and characterization of the collections increase the visibility and usefulness of this valuable germplasm to the stakeholder community. Phenotypic descriptors have great utilization and help guide breeders and others who must decide which accessions will help advance their program. Therefore, phenotyping is a focus of the NCGR. Genotypic descriptors, such as SSR and SNP markers are also developed for the collections and are useful to the scientific community and for combining with rich phenotypic data to discover QTLs. Objective 3 Research Goal 3: Regularly document best management practices and update Crop Vulnerability Statements every 3-4 years. Approach: This objective describes developing updated and documented best management practices for collection management and evaluation. It also is important to have up-to-date crop vulnerability statements to facilitate communication with stakeholders, identify gaps in the collection, and identify management improvements potentially increasing yield and quality of regenerated propagules. In addition, Crop Vulnerability Statements are used by Office of National Programs when communicating with leadership and lawmakers.

Progress Report
Significant progress was made against all project objectives. However, because of a storm in Beltsville, Maryland, during the winter of 2018-2019, the roof blew off the greenhouse with the wild Prunus (almond, apricot, cherry, peach, and plum) seedlings, making retesting for pathogens necessary. Therefore, the seedlings were not received in 2019, and a double batch of about 300 wild Prunus seedlings is expected in 2020. Because of this, no new germplasm was received in FY19. This is acceptable because of a lack of sufficient field space to accommodate large numbers of new plants. Knowledge of amount and patterns of distribution of genetic diversity within and among species gene pools is of considerable value for effective conservation, enrichment, management, and utilization of genetic resources. We use microsatellite markers and single nucleotide polymorphisms to quantify and describe the patterns of distribution genetic diversity, establish the genetic identity of accessions, classify them based on genetic similarities and distances, and analyze genetic structure differentiation. The table and wine grape collections were genetically characterized using 21 microsatellite loci and single nucleotide polymorphisms (SNPs). This project is currently in progress. Upon completion, the data will be analyzed for assessing genetic structure and differentiation in conjunction with the ~1800 SNP loci data. The goal of this project is to compare the effectiveness of microsatellite polymorphisms and SNPs in assessing the genetic diversity and classification of grape germplasm. The DNA for the entire grape collection including cultivated, wild and hybrids is being assembled in a format (96-well plates with standards) required for genotyping ~2000 amplicon-based sequence variation generating multi allelic amplicon polymorphisms with richer information content than SNPs. This is a collaborative project with VitisGen. An ongoing project deals with genotyping 681 pistachio trees in the cultivated and wild species germplasm representing eleven species. We are using 16 microsatellite loci with a goal of genetic identification to aid in germplasm management and to assess genetic diversity in the collection, especially within and among the sects of the genus Pistacia. We have analyzed genetic structure and differentiation within the cultivated walnut using a data set comprising 19 SSR loci and 1300 SNPs, alone and in combination to compare the two marker systems in deciphering the patterns of distribution of genetic diversity in the collection. The results are being written into a publication. Staff also published papers focused on characterization of the collections and species therein. These included discovery of viruses in the pistachio collection and evaluation of pomegranates including growing site, fruit quality, consumer preference, photosynthesis, and chemical content including punicalagins. Researchers published research on the grape collection including characterization of grapevine leafroll-associated virus and genetic diversity of cultivated and wild grapes. They also published research on the fig collection focusing on DNA profiling of the collection along with a collection from Slovenia. In addition, they published research on the persimmon collection focusing on sensory attributes and hedonic rankings of dried fruit and nondestructive determination of astringency. Staff also published papers on an age-related disorder of almonds, a genome sequence for walnut and a study about the robustness of the native northern California black walnut population. Propagation and management of germplasm are within the important objective of maintaining the plants in the collection and working toward no plant mortality by maintaining young, vigorous trees and vines. The almond collection is being imperiled by oak root fungus (Armillaria) and an increasing number of trees are dying each year. This has necessitated repropagation of the entire almond collection for relocation on our land at Wolfskill Experimental Orchard. Because persimmons have resistance to Armillaria, they have also been repropagated. Some grafts of both collections were made this year to complete their propagation. One row of persimmons has been planted beside the almond collection and two and one-half rows of almonds have been planted beside the persimmon collection and their locations are swapped. Trees will need to be removed to accommodate replanting both collections. This is underway. Shading snow fencing was erected to provide protection from the sun to young, cold-hardy kiwifruit that are being added to the collection. The result is that the plants are establishing and growing much better than when they were in full sun. Also woody weeds were removed from the kiwifruit collection. A focus this year was also on pruning, maintenance, and woody weed removal from the walnut, plum, and apricot collections. They are much improved. The grape collection, walnuts, wild peaches, young trees in the nursery rows (newly received material), cherries, olives, pomegranates, and mulberries were all pruned by mechanical hedging. This is necessary to maintain the collections at their current tight spacing. Distributions of National Clonal Germplasm Repository (NCGR) germplasm are primarily winter collected, dormant cuttings or scionwood; although the NCGR also distributes leaves, summer cuttings, pollen, fruit and other plant parts as requested. Almost no seeds are distributed. Because dormant cuttings are primarily distributed, nearly all orders are shipped in late winter/early spring. Each item shipped is three to five cuttings/item (accession). During the past year, 307 orders for germplasm were filled and 2,650 order items (germplasm from individual plants) were shipped, primarily to the research community. Of these 2,650 order items, 2,478 (93.5%) were sent to domestic addresses in the U.S. and 172 (6.5%) order items were shipped abroad. The genetics that were requested are necessary for these scientists to meet their research objectives. There are two copies of clonal accessions and up to 10 seedlings from wild accessions in the collections. These are generally not backed up at another location and represent a vulnerability to the collections and an important objective for the NCGR. Therefore, the NCGR has been working with the National Laboratory for Genetic Resources Preservation (NLGRP) in Fort Collins, Colorado, for cryopreservation studies. Success has been achieved with cryopreservation of pollen of peach and other members of the Prunus genus. Therefore, about 30 accessions are now in cryo-storage. Additionally, wild relatives that self-pollinate can be stored as seeds, and about 12 accessions of wild peach seeds are backed up in cryo-storage. Success has recently been achieved with vegetative shoot tips of grape accessions.

1. New technology changes the conservation status of northern California black walnut. A study was done by ARS researchers in Davis, California, using microsatellite markers to determine the genetic purity status of the northern California black walnut (Juglans hindsii). DNA was extracted from 158 mostly wild J. hindsii trees from 10 counties in northern and southern California and one county in southern Oregon. The study found that at least 71.5% of the wild J. hindsii trees represent pure members of that species. As a result, the California Native Plant Society has reclassified the species so that it is no longer given conservation status. The species is robust.

2. Sequencing and complete assembly of heterozygous genomes of J. regia and J.microcarpa. Members of the genus Juglans are monoecious wind-pollinated trees in the family Juglandaceae with highly heterozygous genomes, which greatly complicates genome sequence assembly. However, ARS researchers in Davis, California, in collaboration with University of California, Davis, Plant Sciences Department, sequenced an interspecific hybrid Juglans microcarpa x J. regia using a novel combination of single-molecule sequencing and optical genome mapping technologies. The resulting assemblies were remarkably complete including chromosome termini and centromere regions. Given the importance of J. microcarpa x J. regia hybrids as potential walnut rootstocks, the researchers catalogued disease resistance genes in the parental genomes and studied their chromosomal distribution. They also estimated the molecular clock rates for woody perennials and deployed them in estimating divergence times of Juglans genomes and those of other woody perennials.

3. Walnut genetic linkage map and QTL analysis of economic traits. A genetic linkage map of walnut containing 2220 single nucleotide polymorphisms (SNPs) in 16 linkage groups (LGs) was constructed by ARS researchers in Davis, California, in collaboration with University of California, Davis, Plant Sciences Department, using an F1 mapping population from a cross between ‘Chandler’ and ‘Idaho’, two contrasting heterozygous parents. Five quantitative yield traits; lateral fruitfulness, harvest date and three nut traits (shell thickness, nut weight, and kernel fill) were then mapped on to the linkage groups. A significant quantitative trait locus (QTL) in LG 11 with negative additive effects suggested heterozygote superiority in the expression of lateral bearing. A set of three QTLs explaining ~10% of the variation in harvest date was located in LG 1. Shell thickness, nut weight, and kernel fill were under the control of two to three linked pleiotropic QTLs in LG 1 segregating from ‘Idaho’. The marginal positive additive effects of QTLs for harvest date, shell thickness, and nut weight and small negative additive effects for kernel fill suggested the QTLs had marginal effect on the expression of these traits.

4. Pistachio genome sequencing and phenotyping. ARS researchers in Davis, California, are collaborating with the University of California, Davis, Genome Center and the Foundation Plant Service on phenotyping and genotyping about 1,000 trees each of grafted and nongrafted hybrid pistachio rootstock seedlings. This is an interspecific hybrid between Pistacia atlantica and P. integerrima that is known as UCB-1. During the previous year, the genomes were sequenced. Recently, the focus was on the phenotypic data and the researchers discovered that early growth is a poor predictor of ultimate growth and vigor of the rootstock. Detailed analysis of the first five years of growth of the UCB-1 seedlings was accomplished.

5. Germplasm backup. The collections at the National Clonal Germplasm Repository (NCGR) in Davis, California, are generally not backed up at another location and represent a vulnerability and an important objective for the NCGR. Therefore, the NCGR has been working with the NLGRP in Fort Collins, Colorado, for cryopreservation studies focused on the Prunus and grapes. Success has been achieved with cryopreservation of pollen of peach and other members of the Prunus genus and with vegetative shoot tips of grape. Pollen was collected from 10 Prunus accessions and shipped to Fort Collins for cryopreservation.

Review Publications
Gradziel, T., Lampinen, B., Preece, J.E. 2019. Propagation from basal epicormic meristems remediates an agring-related disorder in almond clones. Horticulturae. 5(28):1-9.
Diaz-Lara, A., Klaassen, V., Stevens, K., Sudarshana, M.R., Rowhani, A., Maree, H.J., Chooi, K.M., Blouin, A.G., Habili, N., Song, Y., Aram, K., Arnold, K., Cooper, M.L., Wunderlich, L., Battany, M.C., Bettiga, L.J., Smith, R.J., Bester, R., Xiao, H., Meng, B., Preece, J.E., Golino, D., Alrwahnih, M. 2018. Characterization of grapevine leafroll-associated virus 3 genetic variants and application towards RT-qPCR assay design. PLoS One. 13(12):e0208862.
Preece, J.E., Aradhya, M.K. 2019. Temperate nut crops: chestnut, hazelnut, pecan, pistachio, and walnut. In: Greene S., Williams K., Khoury C., Kantar M., Marek L., editors. North American Crop Wild Relatives, Volume 2. Cham, Switzerland: Springer. p. 417-449.
Chater, J.M., Mathon, C., Larive, C.K., Merhaut, D.J., Tinoco, L.W., Mauk, P.A., Preece, J.E. 2019. Juice quality traits, potassium content, and 1H NMR derived metabolites of 14 pomegranate cultivars. Journal of Berry Research. 9(2):209-225.
Aradhya, M.K., Ibrahimov, Z., Toktoraliev, B., Maghradze, D., Musayev, M., Bobokashvili, Z., Velasco, D., Preece, J.E. 2017. Genetic and ecological insights into glacial refugia of walnut (Juglans regia L.). PLoS One. 12(10):e0185974.
Riaz, S., De Lorenzis, G., Velasco, D., Koehmstedt, A., Maghradze, D., Bobokashvili, Z., Musayev, M., Zdunic, G., Laucou, V., Walker, A.M., Failla, O., Preece, J.E., Aradhya, M.K., Arroyo-Garcia, R. 2018. Genetic diversity analysis of cultivated and wild grapevine (Vitis vinifera L.) accessions around the Mediterranean basin and Central Asia. Biomed Central (BMC) Plant Biology. 18:137.
Guzman, F., Segura, S., Aradhya, M.K., Potter, D. 2018. Evaluation of the genetic structure present in natural populations of four subspecies of black cherry (Prunus serotina Ehrh.) from North America using SSR markers. Scientia Horticulturae. 232:206-215.
Milczarek, R.R., Liang, P., Wong, T., Augustine, M.P., Smith, J.L., Woods, R., Sedej, I., Olsen, C.W., Vilches, A.M., Haff, R.P., Preece, J.E., Breksa, A.P. 2019. Nondestructive determination of the astringency of pollination-variant persimmons (Diospyros kaki) using near-infrared (NIR) spectroscopy and nuclear magnetic resonance (NMR) relaxometry. Postharvest Biology and Technology. 149:50-57.