Location: Plant Physiology and Genetics Research2018 Annual Report
1. Develop genotyping-by-sequencing methods for diverse cotton, oilseed, and industrial crop germplasm, and map genetic markers for economically and agronomically important traits in these crops. 1.1 Genotyping-by-sequencing of cotton. 1.2 Genotyping-by-sequencing of oilseed rape. 1.3 Genetic and phenotypic characterization of new guayule germplasm. 2. Develop novel phenotyping approaches for quantitative genetic analysis of drought and heat tolerance traits in cotton and oilseed crops. 2.1 High-throughput phenotyping of traits related to drought and heat tolerance in cotton. 2.2 High-throughput phenotyping of traits related to drought and heat tolerance in oilseed rape. 3. Identify molecular markers associated with stress tolerance traits in cotton and oilseed crops. 3.1 Genome-wide association studies and marker-trait validation in cotton. 3.2 Genome-wide association studies and marker-trait validation in oilseed rape.
The three objectives of the plan will be carried out using various field-based instrumentation and modern approaches in plant breeding and genetics, and will be focused on the three important crops; cotton, oilseed rape, and guayule. These approaches are targeted toward the creation of genotyping-by-sequencing marker maps, development of novel phenotyping tools for quantitative genetic analysis of heat and drought tolerance traits, and identification of molecular markers associated with stress tolerance, quality, and yield related traits. The experiments will apply translational genomics approaches, leveraging statistical genetics and genomics for dissection of quantitative traits and the utilization of rapid, high-throughput phenotyping.
This is the final report for this project which terminated June 2018. Please see the report for the new replacement project, 2020-21410-007-00D, “Enhancing Abiotic Stress Tolerance of Cotton, Oilseeds, and Other Industrial and Biofuel Crops Using High Throughput Phenotyping and Other Genetic Approaches”, for additional information. Substantial results have been realized over the 5 years of the project. Objective 1 focused on development of genotyping-by-sequencing (GBS) methods and the associated databases for diverse cotton, oilseed, and industrial crop germplasm, for subsequent use in genome-wide association studies (GWAS) to identify alleles/genetic markers for economically and agronomically important traits in these crops. GBS methods were selected since they provide large amounts of DNA sequence information and can be used to characterize population structure and diversity. In collaboration with scientists at Cornell University, Ithaca, New York, a diverse collection of approximately 800 Brassica napus genotypes was analyzed using GBS. Comparison of the DNA sequences between these various genotypes allowed for the identification of approximately 30,000 informative molecular markers. Analysis of these markers helped define three distinct and diverse groups of plants that could be distinguished by growth habit and geographical origin including spring, winter-European and winter-Asian subgroups. This work provided insight to the diversity of genotypes available within the B. napus population and provides molecular markers that can be used to identify genes and genomic regions that are associated with traits of interest. GBS technologies were also recently applied to guayule, which is a natural source of rubber. Understanding the genetic diversity in guayule is essential for increasing rubber yields and stress tolerance using molecular breeding approaches. In collaboration with scientists at Cornell University, Ithaca, New York, GBS data were obtained for the entire USDA guayule germplasm collection, as well as several closely related plant species. The GBS analyses identified approximately 50,000 informative molecular markers. Analysis of the relationships between these markers provided significant insight to the evolutionary history of the plant population. The degree of genetic diversity among the guayule accessions (domesticated, breeding lines and wild) was also informative as to which genotypes are most useful for future breeding efforts to improve rubber and other traits. This information underpins future studies to determine the entire genome sequence of one of the guayule lines. A draft genome sequence was developed as a result of collaborative work between scientists at Cornell University, Ithaca, New York, ARS labs in Albany, California, and Maricopa, Arizona, the International Maize and Wheat Improvement Center (CIMMYT), Nairobi, Kenya, Kansas State University; Manhattan, Kansas and West Virginia University, Morgantown, West Virginia. Availability of GBS data and a full genome sequence will undoubtedly accelerate breeding efforts to select plants with higher rubber content and better response to changing environmental conditions. GBS technology was also applied to the analysis of Camelina sativa, an emerging oilseed crop. In collaborative research between scientists at the ARS lab in Maricopa, Arizona, and the Donald Danforth Plant Science Center in St. Louis, Missouri, a total of 7,000 high-quality molecular markers were identified using GBS technology in the core collection of spring Camelina sativa accessions. Further characterization of the genetic diversity and population structure was used to infer how plant breeding and selection may have affected the formation and differentiation within Camelina natural populations and how their genetic diversity can be used in future breeding efforts. Our findings provide important information to enhance genetic gain in Camelina breeding programs by allele/gene identification using GWAS and marker-assisted selection. The GBS data will be the core of Objective 2 of the new project to identify and develop molecular markers associated with stress resistance in these crops. Objective 2 was to develop novel field-based, high-throughput phenotyping (FB-HTP) approaches for cotton and oilseed crops. Several phenotyping platforms were established, and a data pipeline was developed. Both FB-HTP and conventional phenotyping methods were used to characterize the variation in heat and drought stress traits in a population of cotton lines provided by collaborators from Arkansas, South Carolina, and the USDA Cotton Germplasm repository in Texas. The trial included two planting dates and four water treatments with three replicates per water treatment. Plant growth was monitored weekly by hand measurements and included soil moisture, growth stage, leaf chlorophyll content, flowering date, photosynthesis, and pollen sterility. Complementary data on plant height, canopy temperature, and biomass were recorded weekly using the FB-HTP tractor. At harvest, the plots were assessed for yield and fiber quality in response to planting and irrigation treatments. The planting dates identified traits heavily influenced by heat stress as opposed to traits influenced by water stress. Understanding these differences allows for better identification of targets for genetic mapping and plant breeding. The second year of this trial was completed in November 2017 and data are undergoing analysis in preparation for publication of the findings. Preliminary results from this study lead to a re-evaluation of the sensors and deployment protocols for field-based high-throughput phenotyping of cotton at this location. Under Sub-objective 2.1 (GWAS and marker trait validation in cotton), a cotton association mapping population consisting of 400 lines, developed at North Carolina State University, Raleigh, North Carolina, was planted and analyzed in Maricopa, Arizona. This trial was led by a collaborator from the University of Arizona, Maricopa, Arizona, and consisted of single-row, 15 foot long, un-replicated plots. The population is being assessed for heat stress with weekly measurements by the FB-HTP platform and unmanned aerial system-based imagery. At harvest, plots will be assessed for fiber quality using the high-volume instrument located at Cotton Inc., Cary, North Carolina. Seed will be kept from this year’s trial and planted again in a replicated design the following year for further assessment and GWAS mapping of heat tolerant traits. These studies are continued in the new project. Replicated trials of elite germplasm and commercial cultivars for Brassica napus, B. carianta, B. juncea, B. rapa, Sinapis alba, and Camelina sativa were planted in Maricopa, Arizona, to characterize the basis of phenotypic variation of Brassicas under drought and heat stresses, FB-HTP data were collected on a weekly basis throughout the growing season using a modified spray-rig Lee Avenger tractor. In addition to FB-HTP measurements, traditional morphological and physiological traits including flowering time, plant height and chlorophyll content, were collected for ground-truthing the HTP data. At physiological maturity, plots were harvested for seed yield. Seed samples were collected for analysis of oil content and fatty acid composition. Significant correlations were observed between the ground truth and HTP related data and traits. As such, the established FB-HTP protocol will be used with other traditional traits to study the genetic variation in the oil-crop populations and diversity panels, which will speed up data collection time and increase the data precision and thus shorten the time required to enhance genetic gain. These studies help in identifying and developing molecular markers associated with stress resistance in cotton and oil crops as outlined in the new project. Objective 3 focuses on identification of molecular markers/candidate genes associated with stress tolerance traits in crops. In collaboration with ARS researchers at Maricopa, Arizona, Peoria, Illinois, Morris, Minnesota, Sidney, Montana, Mandan, North Dakota, Temple, Texas, Ames, Iowa, Akron, Colorado, Pendleton, Oregon, and scientists at Idaho State University, Pocatello, Idaho and Cornell University, Ithaca, New York, grew, over two years, a spring Brassica subset containing 240 accessions in replicated field trials at Akron, Colorado, Ames, Iowa, and Genesee, Idaho. GWAS was conducted for fatty acid profiles to discover useful alleles/genes controlling biofuel traits. Initial results showed associations between fatty acid conversion efficiency and single nucleotide polymorphism (SNP) markers in the vicinity of fatty acid biosynthetic genes, fatty acid desaturase 3 (FAD3) and 3-ketoacyl-CoA synthase 18 (FAE1). This work is ultimately leading to the identification of SNP markers for genomics-assisted breeding approaches to increase the genetic gains of traits related to biofuel production in Brassica breeding programs. Additional efforts focused on studying leaf cuticle wax content and composition in Camelina, which could directly affect the rate of leaf water loss and thus the susceptibility of plants to drought conditions. In collaboration with scientists at West Virginia University, Morgantown, West Virginia, ARS researchers at Maricopa, Arizona, planted a core collection of spring Camelina types and characterized leaf wax content and composition. A GWAS analysis revealed significant molecular markers that were putatively associated with wax-related traits. The putative functions of these markers can be clustered into three major categories including peroxisome movement, defense in pathogen attack, and repression of photomorphogenesis. These findings could provide the basis for future breeding efforts to develop Camelina varieties with superior abiotic resistance traits.
1. Characterization of genetic diversity in a USDA guayule germplasm collection. Guayule (Parthenium argentatum) is a woody perennial shrub native to the desert regions of northern Mexico and southwestern United States that produces natural rubber in its bark tissues. Attempts to increase rubber yields through crop breeding, however, have been hampered by a lack of well-characterized germplasm. ARS scientists in Maricopa, Arizona, along with scientists at Cornell University, Ithaca, New York, and West Virginia University, Morgantown, West Virginia, performed a detailed assessment of all publicly available guayule germplasm, including closely related species and interspecific hybrids. By using a combination of next generation sequencing technologies and phylogenetic approaches, the scientists could clearly determine the genetic identity and relationships for each accession. Overall, these data help to identify specific lines that can be used for crop breeding, identify geographical regions that should be explored to obtain additional genetic diversity, and provide robust molecular tools to enable genomics-assisted crop improvement. These newly developed methods represent a substantial step forward in the development of guayule as an alternative, commercial source of natural rubber.
2. Characterization of leaf waxes in a domesticated Camelina panel. The leaf cuticle contains a waxy protective layer that has low permeability to water, which directly affects the rate of leaf water loss and thus the susceptibility of plants to drought conditions. Leaf waxes were characterized in the Camelina diversity panel. ARS scientists in Maricopa, Arizona, in collaboration with researchers at West Virginia University, Morgantown, West Virginia, analyzed the phenotypic variations within domesticated Camelina using a core collection of 213 spring-type accessions which originated from Europe and North America. The diversity panel exhibited a wide range in total leaf wax contents, wax classes and constituents. Newly developed methods in Camelina core collection provide important information for the alleles/genes that are putatively controlling wax-related traits and will be useful in future breeding efforts to help select the Camelina varieties with superior abiotic resistance traits.
Ilut, D.C., Sanchez, P.L., Coffelt, T.A., Dyer, J.M., Jenks, M.A., Gore, M.A. 2017. A century of guayule: Comprehensive genetic characterization of the US national guayule (Parthenium argentatum A. Gray) germplasm collection. Industrial Crops and Products. 109:300-309.
Thompson, A.L., Thorp, K.R., Andrade-Sanchez, P., Conley, M.M., Heun, J.T., Dyer, J.M., White, J.W. 2018. Deploying a proximal sensing cart to identify drought-adaptive traits in upland cotton for high-throughput phenotyping. Frontiers in Plant Science. 9:507. https://doi.org/10.3389/fpls.2018.00507.
Tomasi, P., Dyer, J.M., Jenks, M.A., Abdel-Haleem, H.A. 2017. Phenotypic variations in leaf cuticular wax classes and constituents in a spring Camelina sativa diversity panel. Industrial Crops and Products. 112:247-251.
Abdel-Haleem, H.A., Foster, M., Ray, D., Coffelt, T.A. 2018. Phenotypic variations, heritability and correlations in dry biomass, rubber and resin production among guayule germplasm. Industrial Crops and Products. 112:691-697. https://doi.org/10.1016/j.indcrop.2017.12.072.
Thompson, A.L., Conrad, A.V., Conley, M.M., Shrock, H., Taft, B., Miksch, C.L., Mills, T.V., Dyer, J.M. 2018. Professor: A motorized field-based phenotyping cart. HardwareX. https://doi.org/10.1016/j.ohx.2018.e00025.