Research Project: Conservation, Characterization, Evaluation, and Distribution of Grain, Oilseed, Vegetable, Subtropical and Tropical Legume, and Warm Season Grass Genetic Resources and Associated Information

Location: Plant Genetic Resources Conservation Unit

2019 Annual Report

Objectives
1. Efficiently and effectively acquire, distribute, and maintain the safety, genetic integrity, health, and viability of priority grain, oilseed, vegetable, subtropical and tropical legume, and warm season grass genetic resources and associated information. 1.A. Acquire genetic resources to expand the diversity of priority crops and crop wild relatives (CWR) available from the genebank via collection, exchange or other appropriate means. 1.B. Conserve and maintain over 94,000 accessions of priority genetic resources and their associated information, periodically assess these priority genetic resources for viability, trueness to type, and health, and distribute accessions upon request. 1.C. Conduct field and greenhouse regenerations of priority crops and CWR to replenish and safeguard high quality genetic resources in state-of-the-art genebank. 2. Develop more effective genetic resource maintenance, evaluation, or characterization methods and apply them to priority grain, oilseed, vegetable, subtropical and tropical legume, and warm season grass genetic resources. Record and disseminate evaluation and characterization data via GRIN-Global and other data sources. 2.A. Using phenotypic descriptors, evaluate priority crops and CWR for agronomic and horticultural traits and incorporate this data into GRIN-Global. 2.B. Develop and apply nuclear magnetic resonance (NMR), rapid N exceed [nitrogen/protein] analyzer (RNEA), high performance liquid chromatography (HPLC), gas chromatography (GC), and gas chromatography-mass spectrometry (GC/MS) procedures to evaluate variation in oil, protein, sugar content, amino acid composition, fatty acid composition, flavonoids, flavors, and other key phytochemicals in priority crops and CWR and incorporate this data into GRIN-Global. 2.C. Develop and apply DNA markers to assess phylogenetic relationships, genetic diversity, population structure, and association with phenotypic traits of priority crops and CWR. Enter DNA genetic marker characterization data into GRIN-Global or other databases (such as GenBank). 3. With other NPGS genebanks and Crop Germplasm Committees, develop, update, document, and implement best management practices and Crop Vulnerability Statements for priority grain, oilseed, vegetable, subtropical and tropical legume, and warm season grass genetic resource and information management.

Approach
Curators will acquire plant genetic resources from collection trips, donations, and exchanges with other gene banks and state universities to adequately conserve the range of crop genetic diversity. Seed from each accession maintained in the collection will be preserved in cold storage to optimize long-term seed viability and reduce the frequency of regeneration. Efforts will continue to conduct standard germination tests on the entire range of crop and crop wild relative accessions in the germplasm collection with emphasis on testing new material and retesting select inventories at ten year intervals. Plant genetic resources (seeds, in-vitro cultures, plants, cuttings, corms, and rhizomes) and associated information will be sent to users worldwide in response to requests received by email, internet, phone, and U.S. mail. Accessions with low seed viability, low seed numbers, original seed only, and age of seed will be targeted for regeneration. Curators will observe and collect phenotypic data using descriptors for each of the accessions/crops grown for regeneration or evaluation. Additional descriptors on classification, local adaptability, and other traits of agricultural importance will be recorded as opportunity permits. Valuable biochemical traits such as oil/fatty acid and protein/amino acid content in oil seed crops; flavonoids and anthocyanins in legumes; flavor and resveratrol in peanuts; protein content in Vigna; protein and mineral content in pearl millet seeds; and fruit color and flavor components in pepper (Capsicum spp.) will be collected, analyzed and made available on the Germplasm Resources Information Network (GRIN-Global). Genetic characterization and evaluation of plant germplasm will be conducted. For genetic characterization of little bluestem, sweet potato and pepper, previously published simple sequence repeat (SSR) markers are available and will be utilized as the focus of the research is not on marker development but rather characterization. For peanut and sorghum, where advanced genomic tools are available, single nucleotide polymorphism (SNP) markers will be used for characterization, association analysis, and design of functional DNA markers. Curators will consult with Crop Germplasm Committees (CGCs) to develop, update, document, and implement Best Management Practices (BMPs) and Crop Vulnerability Statements (CVS) for crops conserved in the genebank. All data including passport, regeneration, and characterization data will be submitted electronically to the Information Technology Specialist or Seed Storage Manager and their designated staff for local storage and uploading to the GRIN-Global database.

Progress Report
A total of 100,342 accessions of 1608 plant species representing 282 genera were maintained in the Griffin plant genetic resources collection. Over 85% of these accessions were available for distribution to users and over 97% were backed up for security at a second location. Bulk seed samples for 83,671 accessions were maintained at -18oC for long-term storage with seed of the remaining accessions stored at 4oC. A total of 46,676 seed and clonal accessions were distributed upon request to scientists and educators worldwide in calendar year 2018 and another 17,485 distributed in 2019 as of June 6, 2019. Sorghum, watermelon, and pepper were the most distributed crops. Long-term clonal maintenance of 188 wild peanuts, 435 warm-season grasses, and 96 bamboo accessions was continued in the greenhouse or field. Seven hundred and fifty-five (756) accessions of sweet potato were maintained in vitro (or in the greenhouse) and serially re-cultured, as necessary. Viability testing has been conducted on 86,687 accessions in total. Of those, 2121 seed inventories were pulled for viability testing in 2018 and 2275 so far for 2019. All of these activities ensure that the crop genetic resources at the Griffin location are safeguarded for future use in developing new cultivars and identifying novel traits and uses in our food and fiber crops. A total of 1451 plant samples were pulled for regeneration this season. Accessions of Pepper Mild Mottle Virus (PMMV) infected Capsicum spp. were increased in the greenhouse for production of virus free seed. Accessions of watermelon, pepper, and cucurbits were regenerated in the field and greenhouse in Griffin, Georgia. Accessions of various vegetable crops were regenerated and phenotyped in collaboration with USDA/ARS Parlier, California, USDA/ARS Mayaguez, Puerto Rico, Rijk Zwaan, HMClause, Bayer, and the World Vegetable Center. Thirty little bluestem accessions from recent collection trips were regenerated in Byron, Georgia and 435 clonal grasses are successfully regenerating in the greenhouse and field plots. Peanut accessions with low seed inventories were planted in Byron and a total of 411 accessions were shelled, cleaned, and submitted to the seed storage laboratory for processing to add to the national collection. A total of 77 newly regenerated accessions of 17 wild peanut species were submitted to the seed storage. Long-term clonal maintenance of the perennial peanut species, Arachis glabrata, and perennial peanut hybrids was continued in the greenhouses. These peanut genetic resources provide a valuable source of variability for plant breeders to use in the development of improved varieties. For seed regeneration of sweet potato wild species, five Ipomoea wild species accessions (PI 538280, PI 540726, PI 543822, PI 543831, and PI 561543) were planted in the greenhouses at Griffin and Puerto Rico. So far, over 850 seeds have been harvested at Griffin. Viability tests were also conducted on 285 lines of morning glory that had been stored over 30 years. Among them, 116 lines had over 10% viability. Based on this data, several of these sweet potato wild species will be added to the germplasm collection and made available for use in sweet potato breeding programs. The genetic relationship among certain of the cultivated species of Capsicum (pepper) remains unclear. This has hindered the development of a Capsicum core collection. Overlapping phenotypic characteristics have thwarted efforts to definitively assign all observed phenotypes to a specific taxon. Collaborative studies continue in an effort to develop and apply Next Generation Sequencing (NGS) techniques to define the boundaries (if they exist) of individual taxa. Many new species of Capsicum (pepper) have recently been described. However, much remains to be determined regarding the genomic relationships between these new identified species and the cultivated species. Work continues to identify and characterize unique genomic components of both. Approximately 200 accessions of pepper were genotyped for eventual selection of a core collection that will facilitate use of the germplasm collection by plant breeders. All species of Citrullus (watermelon) are monoecious – except one. The species of Citrullus most distantly related to the cultivated watermelon is dioecious, indicating an ancient shift from dioecy to monoecy. Work continues in an effort to identify the genetic/genomic basis for this shift as part of a broader pan-genomic characterization of the genus. Collaboration continues with ARS Charleston and the Cucurbit Coordinated Agricultural Project (CucCAP) regarding genotyping and phenotyping of Citrullus germplasm. Evaluation of the basis for insect resistance in the desert perennial vine (Citrullus ecirrhosus spp.), a relative of cultivated watermelon, continues in collaboration with USDA/ARS Oxford, Mississippi and USDA/ARS Charleston, South Carolina. Genomic studies in Citrullus (sex determination, general genomics). In a set of 38 cowpea accessions evaluated for biochemical and seed traits, the main components of variation within this set was attributed to anthocyanins, flavonols, 100 seed weight, seed pattern, and seed pattern color. Cluster analysis grouped the 38 cowpea accessions into five anthocyanin and flavonol groups. A group of 27 roselle accessions which were previously thought to be non-seed producers because of photo-period sensitivity were tested for viability. Percent viable ranged from 0 to over 80% for these roselle accessions and several exceeded 65%. The information obtained in these studies will provide plant breeders and scientists valuable biochemical variability data in cowpea and germination data in roselle for development of important cultivars as well as the identification of roselle genotypes which can produce high quality seed quantities. Fifty-two high oleic acid peanut accessions were grown in replicated field trials at three locations (Georgia, Florida, and New Mexico). Seeds were harvested and used for oil, fatty acid, and protein analysis. The 52 peanut lines were genotyped with FAD2A and FAD2B DNA markers. These DNA markers were developed to easily identify high oleic acid peanut varieties. The interaction between the different FAD2A and FAD2B genotypes with the environment was found to be significant and could alter the amount of oleic acid produced. The revealed interaction will be very useful to peanut breeders, farmers, and processors of peanut products for understanding and utilizing this important trait. In a separate study, 14 high stearic acid (C18:0) peanut accessions were planted in the field trials to determine if levels of stearic acid previously measured in long-term storage peanut samples would be consistent in newly harvested seed. Significant differences between the freshly harvested and long term-stored peanut seed were identified. Additional germplasm characterizations performed at the location include 146 samples (104 cultivated peanut, 19 Desmodium, 9 lablab, and 12 Corchorus) measured for protein content and 44 samples (19 Desmodium and 25 Vigna) measured for flavonoid content. In collaboration with the University of Georgia and ARS scientists in Lubbock, Texas, 256 EMS-induced mutant sorghum lines were evaluated for low-phosphorous tolerance at the lab conditions. Ten high oleic acid sesame lines (M5-M6) were planted in the greenhouse in 2018 to confirm the high oleic trait identified in previous studies.

Accomplishments
1. High oleate is a health beneficial seed quality trait frequently incorporated in peanut varieties. Crop wild relatives are useful in crop improvement, but for wild peanut species, many chemical or nutritional traits are not well characterized. Using gas chromatography, a group of 209 populations of 45 wild peanut species maintained by the USDA National Plant Germplasm System was screened for the high oleate trait. A naturally occurring mutant from the wild peanut species, Arachis veigae S. H. Santana & Valls, with increased oleic and very long chain fatty acid content was identified. Using DNA sequencing, ARS researchers in Griffin, Georgia, discovered a genetic mutation which may be responsible for the trait by reducing enzyme activity in the oleic acid pathway. This discovery will improve peanut breeding programs by providing a source of the high oleate trait in wild species while enhancing the genetic diversity in the crop. Using this wild species as a parent in a breeding program allows for incorporation of this trait along with other novel traits found in peanut crop wild relatives such as disease resistance and drought tolerance.

Review Publications
Jarret, R.L., Barboza, G., Batista, F., Chou, Y., Hulse-Kemp, A.M., Ochoa-Alejo, N., Veres, A., Berke, T., Carrizo, C., Csillery, G., Huang, Y., Kiss, E., Kovacs, Z., Kondrak, M., Arce-Rodriguez, M., Scaldaferro, M.A., Szoke, A., Tripodi, P. 2019. Capsicum - an abbreviated compendium. Journal of the American Society for Horticultural Science. 144(1):3-22. https://doi.org/10.21273/JASHS04446-18.
Tonnis, B.D., Wang, M.L., Tallury, S.P., Tishchenko, V., Stalker, H. 2019. Identification of a mutant from Arachis veigae with enhanced seed Oleic and very long chain acid content. Applied Biological Chemistry. 62:9. https://doi.org/10.1186/s13765-019-0420-x.
Srinivasan, R., Abney, M., Lai, P., Culbreath, A., Tallury, S.P., Leal-Bertioli, S. 2018. Resistance to Thrips in peanut and implications for management of Thrips and Thrips-transmitted Orthotospoviruses in peanut. Frontiers in Plant Science. 9:1604. https://doi.org/10.3389/fpls.2018.01604.
Hancock, W., Tallury, S.P., Isleib, T., Chu, Y., Ozias-Akins, P., Stalker, T. 2019. Introgression analysis and morphological characterization of an Arachis hypogaea × A. diogoi interspecific hybrid derived population. Crop Science. 59(2):640-649. https://doi.org/10.2135/cropsci2018.07.0461.
Morris, J.B., Wang, M.L. 2018. Updated review of potential medicinal genetic resources in the USDA, ARS, PGRCU industrial and legume crop germplasm collections. Industrial Crops and Products. 123:470-479. https://doi.org/10.1016/j.indcrop.2018.07.014.
Wu, S., Wang, X., Reddy, U., Sun, H., Bao, K., Patel, T., Oritz, C., Abburi, L., Nimmakayala, P., Branham, S., Wechter, W.P., Massey, L.M., Ling, K., Kousik, C.S., Hammar, S.A., Tadmor, Y., Portnoy, V., Gur, A., Katzir, N., Guner, N., Davis, A., Hernandez, A.G., Wright, C.L., McGregor, C., Jarret, R.L., Xu, Y., Zhang, X., Wehner, T.C., Grumet, R., Levi, A., Fei, Z. 2019. Genome of ‘Charleston Gray’, the principal American watermelon cultivar, and genetic characterization of 1,365 accessions in the U.S. National Plant Germplasm System watermelon collection. Plant Biotechnology Journal. https://doi.org/10.1111/pbi.13136.
Jing, L., Yueyi, T., Jacobson, A., Dang, P.M., Wang, M.L., Hagan, A., Chen, C. 2018. Population structure and association mapping to detect QTL controlling tomato spotted wilt virus resistance in cultivated peanuts. The Crop Journal. https://doi.org/10.1016/j.cj.2018.04.001.
Wang, L., Zhang, Y., Li, D., Dossa, K., Wang, M.L., Zhou, R., Yu, J., Zhang, X. 2019. Gene expression profiles that shape high and low oil content sesames. BMC Genetics. 20:45. https://doi.org/10.1186/s12863-019-0747-7.
Wadl, P.A., Olukolu, B.A., Branham, S., Jarret, R.L., Yencho, G., Jackson, D. 2018. Genetic diversity and population structure of the USDA sweetpotato (Ipomoea batatas) germplasm collection using GBSpoly. Frontiers in Plant Science. 9:1166. https://doi.org/10.3389/fpls.2018.01166.
Jackson, D., Harrison, H.F., Jarret, R.L., Wadl, P.A. 2019. Phenotypic analysis of leaf colours from the USDA, ARS sweetpotato (Ipomoea batatas) germplasm collection. Plant Genetic Resources. https://doi.org/10.1017/S1479262119000042.
Morris, J.B., Wang, M.L., Tonnis, B.D. 2019. Variability for Sennoside A and B concentrations in eight Senna species. Industrial Crops and Products. 139:111489. https://doi.org/10.1016/j.indcrop.2019.111489.