Research Project: Exploiting Genetic Diversity through Genomics, Plant Physiology, and Plant Breeding to Increase Competitiveness of U.S. Soybeans in Global Markets

Location: Soybean and Nitrogen Fixation Research

2023 Annual Report

Objectives
Objective 1: Use genomics, physiology and plant breeding approaches to identify novel genetic variation for various yield-enhancing traits in the USDA germplasm collection, transfer the traits to adapted backgrounds, and release germplasm or cultivars with improved yield potential. Sub-obj. 1a: Identify desirable genetic diversity for seed yield in exotic Asian soybean cultivars. Sub-obj. 1b: Identify desirable genetic diversity for improved seed yield in wild soybean. Sub-obj. 1c: Develop improved breeding methods and approaches for incorporation of genetic diversity from wild soybean to applied breeding programs. Sub-obj. 1d: Identify genomic differences between F1 hybrids and inbred parents that can be exploited as the basis for new breeding methodologies to augment existing applied breeding pipelines. Objective 2: Identify and characterize genetic variation for soybean oil and protein in the USDA germplasm collection, transfer the traits to adapted germplasm, and release improved germplasm or cultivars. Sub-obj. 2a: Elucidate physiological processes by which seed composition is improved without a yield penalty and connect these to the underlying genes. Sub-obj. 2b: Introgress desirable combinations of protein genes/QTL into MG V-VII high yielding conventional elite backgrounds. Sub-obj. 2c: Identify new genetic resources with high seed protein that lack the DBK high protein allele on chromosome 20 and identify QTL in those sources. Sub-obj. 2d: Determine if alterations of N metabolism and expression of asparagine synthase (AS) genes in vegetative tissues are associated with improved N content in progeny derived from the backcross of NMS4-44-329 to its parent N7103. Sub-obj. 2e: Determine if variation in a small RNA reported to regulate S uptake in Arabidopsis plays a similar role in soybean and characterize its mechanism of action. Sub-obj. 2f: Introgress desirable combinations of oil genes/QTL into MG V-VII high yielding conventional elite background to develop and release high performing germplasm with improved oil quantity and quality. Sub-obj. 2g: Combine high seed protein with drought tolerance in high yielding soybean backgrounds. Sub-obj. 2h: Improve soybean seed oil content and composition using wild soybean. Objective 3: Screen the USDA soybean germplasm collection to discover and elucidate traits governing genetic variation for molecular and physiological mechanisms that preserve yield under dry conditions, and use the information to develop and release soybeans with improved drought and heat tolerance. Sub-obj. 3a: Develop adapted drought tolerant breeding lines from exotic soybean germplasm. Sub-obj. 3b: Quantify the yield impact of the slow-wilting trait on yield in diverse environments. Sub-obj. 3c: Determine the impact of limited transpiration on leaf gas exchange and seed yield during drought. Sub-obj. 3d: Identify physiological and molecular traits that underlie successful nitrogen fixation response to drought. Sub-obj. 3e: Determine the ability of wild soybean accessions to germinate and grow at suboptimal temperatures and identify inheritance.

Approach
The USDA Soybean Germplasm Collection is one of the greatest biological resources in the world and a premier source of new genes for key soybean traits. Our team of experts genetically mines the Collection through breeding, genomics and plant physiology to provide novel customer-ready breeding stocks and production know-how to the soybean industry and society. The three objectives use wild and domesticated soybean germplasm from around the globe as a genetic basis for improving the yield potential and economic value of the U.S. soybean crop, while protecting crop production from the ravages of weather extremes, especially drought. A common approach in all three objectives is to blend cutting-edge field and lab research to transfer novel alleles and traits from the Collection into adapted, high-yielding, publicly-available USDA cultivars and breeding lines. Innovative plant breeding teams up with physiological and genomic research to make breeding advances and determine the mechanistic and molecular basis for them. These discoveries guide and refine future mining of the Collection, improving overall efficiency in utilizing the Collection and amplifying its impact. Because more than 90% of U.S. soybean acreage is grown in private varieties, private industry, rather than the public sector, will be the most immediate user of the novel USDA-ARS breeding stocks developed in this project. However, because all our products are non-GMO, they will also be used directly in the small but fast-growing conventional and organic soybean markets. To ensure the successful transfer of USDA products to the farm (either directly or more indirectly as breeding stock for private industry) it is essential that our USDA-ARS germplasm releases be ‘user ready’. In other words, germplasm released from the project must yield within 5% of current commercial cultivars, deliver stable traits and, when possible, include associated genetic markers. Integration of genomics, molecular biology and plant physiology with the top-notch ARS field breeding program makes this goal achievable.

Progress Report
This is the final report for this project. Soybean is one of the least genetically diverse crops in the USA. As a result, it lacks the genetic capacity to resist emerging threats from new pests and weather extremes or adapt to new market demands. U.S. soybean breeders do not have the genetic resources in their programs to breed to combat these problems and increase the sustainability and profitability of US soy. The USDA Soybean Germplasm Collection (SGC) includes over 20,000 domesticated (Glycine max) and wild (Glycine soja) accessions that represent most of the global diversity of soybean and is a vital resource to increase genetic diversity. The Soybean and Nitrogen Fixation Research Unit (SNFRU) continues to identify economically important genes and alleles preserved among the USDA SGC and transfer them into user-friendly germplasm, which are easily accessible to public and private sector scientists throughout the U.S. Soybean meal (seed residue after oil extraction) is the world’s premier high-protein feed source for livestock and accounts for 60-70% of the total value of soybean. However, the protein content of meal produced from the majority of recently released commercial U.S. soybean cultivars has dropped below a global standard of 47.5%. A decline that has been attributed to a pervasive negative correlation between seed protein content and yield, which until recently was largely neglected. Additionally, soymeal is deficient in 5 important amino acids that must be added as supplements when the soymeal is fed to animals. The SNFRU is addressing these problems by identifying and utilizing both wild and domesticated accessions as breeding stocks and by evaluating the role of agronomic practices in improving soy protein. PROJECT SUMMARY of 6070-21220-069-000D entitled “Exploiting Genetic Diversity through Genomics, Plant Physiology, and Plant Breeding to Increase Competitiveness of U.S. Soybeans in Global Markets” that ended in March 2023. Lack of genetic diversity is one of the top concerns of the U.S. soybean community. We report success in increasing genetic diversity, especially in increasing genetic diversity through hybridization of cultivated (G. max) with wild soybean (G. Soja) without reducing seed yield. Prior to this research wild soybean was considered a poor choice for soybean improvement even though it is more genetically diverse than the G. max germplasm because the F2 progeny looks distinctly poor. We have developed more than a dozen breeding lines with 12.5-50% soja genome that are highly competitive with commercial check cultivars in seed yield, seed composition and overall agronomic desirability. Through regional testing 12 and 6 lines were identified that yielded >95% of the test mean that contain a 12.5% and 25% pedigree derived from the SGC, respectively. Many of these lines also exhibited meal protein contents >48%. Therefore, progress is being made to develop breeding lines from wild soybean, that are genetically diverse with high yields and improved seed composition. The correlation between seed protein content and yield was thought to be intractable, however breeding with diverse genetic resources produced progenies with high yield and high seed protein. Since 2020, we have released three germplasm (USDA-N6005, USDA-N7004, USDA-N5001) with 2-3% higher meal protein and same or better yield than the check cultivars. Drought is one of the biggest concerns in sustainable agriculture. It is predicted that in the coming years there will likely be an increase in the frequency and intensity of droughts due to a changing climate. The Unit has released one germplasm line (USDA-N7006) with drought tolerance and co-released two more drought tolerant germplasm lines within the last five years. Under drought stress these lines exhibit lower canopy wilting and a yield increase of >10-20%. Two slow canopy wilting lines were shown to have an increased water use efficiency (WUE) and ozone tolerance. WUE of the two breeding lines increased by as much as 30% when compared to other slow canopy wilting lines. Foliar injury from ozone damage decreased by as much as 75% when compared to sensitive checks. To improve screening strategies for drought tolerance, it is critical to understand physiological mechanisms that are associated with drought tolerance. The unit has examined the physiology of several soybean genotypes under well-watered and water-limited conditions. These studies have included genotypes previously identified by breeders as drought tolerant or susceptible and genotypes whose drought response physiology has been investigated in growth chamber studies. Results indicate that drought response mechanisms differ depending on the severity of drought stress, and physiological traits that are expected to confer drought tolerance may not translate to yield resilience during drought in some genotypes. The findings also indicate that growth chamber studies are not always reliable predictors of drought response mechanisms in the field, highlighting the importance of field-based research to complement controlled environment studies. The yield loss from flooding can be just as detrimental as drought, causing significant economic loss. Over the last 40 years, flooding alone has cost the U.S. an estimated $161.6 billion in damages. The results collected from the past five years demonstrate we have been successful in breeding for flood tolerance in soybean. Breeding lines with the highest level of flood tolerance have been shown to continually outperform susceptible genotypes, when grown under flooded conditions, with an average 25% higher seed yield. Over the last 5 years, we published 49 research articles in refereed journals and released eight high yielding germplasm with increased genetic diversity, improved seed composition, or drought tolerance. our research was augmented by over $11.8M in extramural funds. The unit SYs have served as PI on more than 20 national and international grants funded by United Soybean Board, North Carolina Soybean Producers Association, and Foundation for Food and Agriculture Research to conduct research to improve genetic diversity, meal protein, drought tolerance, and test weight. 2023 PROGRESS REPORT. The Unit grew over 80 acres of soybean to evaluate yield and produce new breeding lines. In 2023 we authored or co-authored 14 publications. SNFRU harvested more than 7000 yield plots and 2500 progeny rows of breeding lines developed from crosses with unimproved Asian or wild soybean accessions. More than 70 elite breeding lines from this research were entered in USDA Regional and the USB Regional Southern Diversity yield trials. Our advanced USDA germplasm derived from unimproved PIs from the SGC is being used by public and private breeders to develop new cultivars with high yields and serve as checks in state and regional yield trials. One germplasm (USDA-N7005) was released that included Asian soybean in their pedigree and a new source of yield-enhancing genes. We are also investigating the impact of management practices, such as fertilization and tillage, on seed protein. Heat stress is often overlooked in breeding programs, in part because symptoms of heat stress can be more subtle than those of drought stress. The unit conducted multiple years of an open-air, elevated temperature field experiment and found significant variation among varieties in physiological responses to heat stress as well as seed composition. These field-based approaches are essential in separating drought effects from heat effects, which are often confounded. We have also begun to identify proteins that regulate temperature-response signaling and outcomes within the plants. These findings will help develop strategies to improve soybean heat tolerance through breeding and biotechnology. The unit coordinates the USDA-ARS soybean winter-nursery in collaboration with USDA colleagues at Mayaguez Puerto Rico facilitating rapid advance for breeding lines. Each year the unit participates in the USDA Southern Uniform Soybean Test (SUST) and coordinates the Protein Diversity Test (PDT) funded by the United Soybean Board. Between these two tests over 300 entries are evaluated in over 1500 plots in two or three locations. The results from each test provide crucial information for fast development and release of new soybean germplasm and varieties.

Accomplishments
1. Released a high yielding maturity group V germplasm USDA-N5001 with high seed and meal protein contents. Soybean stake holders are extremely concerned about low meal protein contents of recent U.S. soybean cultivars that are below the minimum market standard of 47.5%. To stay competitive in the international market, U.S. soybean growers and processors require high-yielding soybean with >48% meal protein. However, increasing protein content without reducing yield is a challenging task due to negative correction of soybean protein and yield. The Soybean Research Unit at Raleigh, North Carolina, has recently released USDA-N5001, combining with 2-3% higher meal protein and same or better yield than check cultivars. This line will be a valuable resource for both public and private soybean breeders to develop high performing soybean cultivars with >48% meal protein. This germplasm was published in Journal of Plant Registrations in July 2023.

2. Breast meat from chickens fed high oleic soymeal have elevated content of oleic acid. Cardiac health benefits are associated with increased oleic acid. Researchers in Raleigh, North Carolina, fed broilers diets that included soymeal with normal levels of oleic acid and elevated levels of oleic acid derived from high and normal oleic near isolines of soybean developed by USDA ARS soybean breeders. Evaluation of the breast meat from the broilers showed that the quality of the meat was high and that broilers fed soy meal with elevated oleic acid content had increased oleic acid content in the meat. High oleic acid soybeans were bred to provide the market with a soy oil that has a stable shelf life with the health benefits of increased oleic acid. Meal from these same lines fed to layers also increase the healthy oleic acid content in the egg.

3. Parental choice and seed size selection improve outcomes when breeding with wild soybean. Many decades of U.S. breeding among few elite parents have created a genetic bottleneck resulting in a very narrow genetic diversity among the current U.S. commercial cultivars. Such narrow diversity makes U.S. soybean vulnerable to pests and environmental stresses, recalcitrant to improvements of seed yield and seed composition, and reduce sustainability and profitability of the crop in the long run. The USDA curated germplasm collection preserves wild soybean that are more diverse than domesticated soybean. However, wild soybean is seldom used in soybean breeding programs because the first generations are strikingly poor in appearance. Researchers in Raleigh, North Carolina, have shown that choice of the domesticated parent in an interspecific cross can substantially improve the number of plants deemed agronomically valuable. More agronomically valuable plants can be recovered if larger seeds are selected. In all cases the genetic diversity of the wild parent could be recovered in a few progenies. This method of breeding will make it easier to bring traits from the wild soybean gene pool such as improved seed composition, tolerance to environmental stress and novel pest resistance to the U.S. soybean gene pool.

4. Improving flood tolerance increases resiliency during crucial growth stages. Improving the plants’ ability to respond and adapt to abiotic stress is needed to address future climate scenarios. Flooding can lead to severe damage, as the standing water and water-logged soils deprive plants of the necessary light, oxygen, and carbon dioxide required for growth. Researchers in Raleigh, North Carolina, developed and evaluated breeding lines for the response to flood stress at three critical growth stages for production – germination, early vegetative growth (V1 and V4), and early reproductive growth (R1). The breeding lines with the highest level of flood tolerance maintained a germination rate of >80% after 8hr of flooding, while the germination rates of the susceptible checks were significantly lower ranging from 58%-63%. During early vegetative and reproductive growth the flood-tolerant lines averaged a 30% decrease in foliar damage and a 10% increase in biomass. Flooding is becoming an increasing concern for soybean production in the U.S. due to an increase of intense rainfall events. One, if not the only, sustainable approach to improve performance under flood stress is to develop flood-tolerant soybean lines.

5. Management strategies can improve soybean seed protein concentration. While breeding for high protein soybean varieties holds promise, growers could benefit from immediate options to implement on their farms to produce the high protein soybean meal desired by end users. Previously, little information was available regarding the impact of management strategies on protein in the Southeast U.S. Researchers in Raleigh, North Carolina, conducted field experiments to examine the impacts of tillage and plant density on soybean protein production across multiple high yielding soybean genotypes. While tillage did not impact seed protein, higher planting density led to higher seed protein concentration in two of three Southeastern U.S. environments tested. Additionally, there was not a yield penalty to higher seed protein concentration in the higher density soybeans. This research reveals that growers may be able to adopt strategies to improve seed protein in the Southeast U.S.

Review Publications
Mathers, C., Heitman, J., Huseth, A., Locke, A.M., Osmond, D., Woodley, A. 2023. No-till imparts yield stability and greater cumulative yield under variable weather conditions in the southeastern USA Piedmont. Field Crops Research. https://doi.org/10.1016/j.fcr.2023.108811.
Ethridge, S., Locke, A.M., Everman, W., Jordan, D., Leon, R. 2022. Crop physiological considerations for combining variable density planting to optimize seed costs and weed suppression. Weed Science. https://doi.org/10.1017/wsc.2022.62.
Kakati, J.P., Fallen, B.D., Bridges, W.C., Nararyanan, S.K. 2022. Characterization of a soybean population for germination and seedling root traits under water stress conditions. Agronomy Journal. 12(8). https://doi.org/10.3390/agronomy12081944.
Miller, M., Song, Q., Fallen, B.D., Li, Z. 2023. Genomic prediction of optimal cross combinations to accelerate genetic improvement of soybean (Glycine Max). Crop Science. 14. Article e1171135. https://doi.org/10.3389/fpls.2023.1171135.
Maharjan, P., Rahimi, A., Harding, K.L., Vu, T.C., Malheiros, R., Oviedo, E.R., Mian, R.M., Joseph, M., Dean, L.L., Anderson, K.E., Toomer, O.T. 2023. Effects of full-fat high-oleic soybean meal in layer diets on nutrient digestibility and egg quality parameters of a white laying hen strain. Poultry Science. 102:102486. https://doi.org/10.1016/j.psj.2023.102486.
Mian, R.M., Cunicelli, M.J., Carter, T., Villagarcia, M.R., Fallen, B.D. 2023. Registration of high yielding maturity group V germplasm USDA-N5001 with high seed and meal protein contents. Journal of Plant Registrations. https://doi.org/10.1002/plr2.20306.
Ramanathan, S., Gannon, T., Locke, A.M., Everman, W. 2023. Characterizing atrazine, mesosulfuron-methyl, and topramezone bioavailability in North Carolina soils using greenhouse bioassays. Agrosystems, Geosciences & Environment. https://doi.org/10.1002/agg2.20371.
Dunmire, K.M., Braun, M.B., Evans, C.E., Stark, C.R., Paulk, C.B., Fallen, B.D. 2023. Determining the apparent ileal digestibility of amino acids for soybean meal from select soybean varieties with varying levels of crude protein. Journal of Applied Poultry Research. 32(2). https://doi.org/10.1016/j.japr.2023.100333.
Wilkes, J., Fallen, B.D., Saski, C., Agudelo, P. 2023. Development of SNP molecular markers associated with resistance to reniform nematode in soybean using KASP genotyping. Euphytica. https://doi.org/10.1007/s10681-022-03144-3.
Toomer, O.T., Oviedo, E.R., Muhammad, A., Patino, D., Joseph, M., Frinsko, M., Vu, T.C., Maharjan, P., Fallen, B.D., Mian, R.M. 2023. Current agronomic practices, harvest & post-harvest processing of soybeans (Glycine max)- A review. Agronomy Journal. 13(2):427. https://doi.org/10.3390/agronomy13020427.
Ostezan, A., Prenger, E., Rosso, L., Zhang, B., Stupar, R., Glenn, T., Mian, R.M., Li, Z. 2023. A chromosome 16 deletion conferring a high sucrose phenotype in soybean. Journal of Theoretical and Applied Genetics. https://doi.org/10.1007/s00122-023-04354-6.
Ethridge, S., Chandra, S., Everman, W., Jordan, D., Locke, A.M., Owen, M., Leon, R. 2023. Rapid evolution of competitive ability in giant foxtail (setaria faberi) over 34 years. Weed Science. 71(1):59-68. https://doi.org/10.1017/wsc.2023.1.
Epie, K., Bauer, P., Stone, K.C., Locke, A.M. 2023. Density, not tillage, increases soybean protein concentration in some southeastern US environments. Agronomy Journal. 115(4):1867–1876. https://doi.org/10.1002/agj2.21371.
Taliercio, E.W., Eickholt, D., Read, Q.D., Carter Jr, T.E., Waldeck, N., Fallen, B.D. 2023. Parental choice and seed size impact the uprightness of progeny from interspecific Glycine hybridizations. Crop Science. 63(4):2184-2195. https://doi.org/10.1002/csc2.21015.