Location: Sunflower and Plant Biology Research
2020 Annual Report
Objectives
Objective 1: Identify, at the genome and physiological levels, plant-plant interactions that impact plant growth and lead to crop yield losses, especially crop-weed interactions that occur during the critical weed-free period, and interactions that occur between the different crops inter-planted in relay cropping systems, such as corn, soybeans, or sunflowers relayed with camelina, ryegrass, or canola. [NP304, Component 2, Problem Statement 2A3]
Sub-objective 1.A: Determine the parameters for evaluating the impacts of winter annual cover crops on corn, sunflower, and Amaranthus spp. productivity.
Sub-objective 1.B: Identify physiological and molecular mechanisms that control interactions between cover crops and corn, sunflower, and Amaranthus spp.
Sub-objective 1.C: Evaluate impacts of candidate genes on cover crop-relay crop and cover crop-weed interactions.
Objective 2: Determine the molecular and physiological mechanisms by which winter annual cover crops suppress weeds in northern temperate agroecosystems, and identify genes that will enhance weed suppression in these crops, such as genes associated with weed-tolerance, cover-crop tolerance, and cold hardiness. [NP304, Component 2, Problem Statement 2A3]
Sub-objective 2.A: Identify genetic markers for improving the weed-suppressing trait of winter hardiness in winter canola and/or camelina varieties.
Sub-objective 2.B: Evaluate the weed-suppressing traits of winter-hardy canola and camelina in the field.
Approach
Weeds are major pests of agro-ecosystems that reduce production of the nation’s food, feed, fiber and fuel crops. The industry-adopted practice of rotating crops with engineered tolerance to a limited set of herbicides continues to put selection pressure on the evolution of herbicide-resistant weeds. As part of a holistic and sustainable approach to managing weeds in temperate agro-ecosystems, we propose to identify cover crop-relay crop interactions to enhance relay crop productivity, cover crop-weed interactions to enhance weed suppression, and identify winter-hardy annual cover crops that suppress weeds in relay cropping systems. In this proposal, winter canola will serve a dual purpose as both a cover crop for evaluating weed suppression, and as a surrogate weed for weed-relay crop interactions. Our model relay cropping system consists of inter-seeding a commodity crop (corn or sunflower) into an established cover crop (winter canola) such that their lifecycles overlap. Currently, no winter-hardy annual broadleaf cover crops are economically suited for weed suppression in relay cropping systems in the upper Midwest (UMW) and Northern Great Plains (NGP). Consequently, the objectives of this project are: (1) elucidating regulatory signals and pathways associated with cover crop-relay crop and cover crop-weed interactions that impact plant productivity, and (2) identifying economically suited winter-hardy broadleaf cover crops that suppress weed establishment in inter-seeded relay crops. These objectives will be accomplished through the use of physiological, molecular, and genomic approaches, with the long-term goals of revolutionizing weed management practices.
Progress Report
Objective 1. To determine if crops detect weeds (competitors) through altered light quality, soil soluble chemical signals, and/or volatile signals, we developed and used a greenhouse-based system to investigate crop-weed interactions. Phenological responses of corn and sunflower varieties to different varieties and densities of winter canola and weedy species indicated that below-ground signals have a greater impact on crop yield than above-ground signals, and that some of the crop responses to below-ground signals can be mitigated by addition of activated charcoal to the soil. Additionally, studies to determine the impact of weed-generated above ground (light quality) or below-ground (diffusible chemical) signals on the transcriptomes of sunflower and corn have been completed and analysis of this data will help identify mechanisms underlying the developmental and physiological pathways that are activated when crops detect weeds.
In collaborative research, we studied the transcriptome response of corn to weeds under both field and greenhouse conditions. Results from those studies implicated corn defense signaling in response to weeds and highlighted the probability that salicylic acid and phytochrome signaling are regulators of competitor-induced yield losses. Gene constructs containing a weed-inducible promotor have been produced to block salicylic acid signaling process in corn only when weeds are present to determine if reducing salicylic acid signaling will make corn less responsive to weed interference. This transcriptome analysis also identified two corn genes induced by weed competition, in both greenhouse and field studies, that contain conserved promotor elements that likely control their induced expression in the presence of competitors. Regulatory elements from these competitor-inducible genes will serve as tools to further investigate the signaling mechanisms responsible for their induction. Several gene constructs designed to test the function of putative weed-inducible promoter elements are in progress and will be used to decipher the signaling mechanisms by which weeds/competitors induce the expression of crop genes.
In the case of sunflower grown under greenhouse conditions in a replicated study, 204 genes were consistently differentially expressed in response to weed interference. This work highlighted a role for oxidative stress in response to weed interference in sunflowers, with indications of abscisic acid, ethylene, and light signaling processes involved in regulation of these genes. Further transcriptome analysis to determine the impact of weed interference on sunflower root are ongoing and preliminary results have been presented at the 2020 Annual Plant Biology meeting.
Because Arabidopsis exhibits a classic critical period for weed control and a classic parabolic loss of yield as weed pressure increases, it is also being used as an efficient model system to study plant-plant interactions. We have used this model to test the role of several regulatory and/or signaling genes similar to those implicated in corn and sunflowers, including genes involved in salicylic acid signaling and red-light signaling. Data from this work has been presented at the America Society of Agronomy annual meeting. Because this model system provides a rapid method for functionally testing genes identified in crop-weed interaction studies, it will be useful in our continued investigation of signaling and molecular mechanisms involved in crop-weed and crop-crop interactions.
Objective 2. Winter oilseed crops and cover crops provide ecosystem services including suppression of weed establishment, but few tolerate the freezing temperatures of northern U.S. climates. A collaboration between researchers at USDA-ARS in Fargo, North Dakota, and Kansas State University identified a winter canola breeding population (totaling 413 accessions) for freezing tolerance studies. To identify genetic traits associated with freezing tolerance, genotyping-by-sequencing of the winter canola diversity panel identified 251,575 high quality single-nucleotide polymorphisms (SNPs). Methods developed for conducting freezing studies using state-of-the-art environmental chambers and high-throughput phenotyping confirmed suitable diversity within this germplasm collection. The genotyping and phenotyping data were used to conduct Genome Wide Association Studies (GWAS), which identified 12 chromosomal locations associated with freezing tolerance in canola and more than 25 potential candidate genes within these regions.
Global climate change increases plant vulnerability to freezing damage either because of insufficient cold acclimation or higher incidence of brief warm spells that cause cold acclimated plants to deacclimate. Research confirmed that warm spells as short as 3 days above 10C(50F) can cause deacclimation, leaving crops such as winter canola vulnerable to freezing conditions. ARS researchers identified four chromosomal regions associated with altered deacclimation rates in our winter canola population. Molecular markers associated with these chromosomal regions provide a starting point for breeders to integrate freezing tolerance into elite breeding lines of both winter- and spring-types of canola. Putative gene function and transcriptomics data generated from a subset of winter canola varieties highlighted ten candidate genes within these four chromosomal regions, including several genes known to be responsive to cold or involved in chromatin modifications in Arabidopsis. Among these 10 genes, Arabidopsis lines containing mutations provided evidence that variations in orthologues of Vernalization Independence 3 (VIP3) and Phytochrome Kinase Substrate 4 (PKS4) play a role in deacclimation processes observed in our winter canola population. We have developed a canola plant regeneration protocol for canola transformation and will use this system to confirm if mutations in VIP3 and PKS4 can mitigate deacclimation processes and reduce freezing damage in winter canola.
In contrast to winter canola, winter biotypes of camelina regularly survive the winter conditions experienced in northern regions of the U.S. and, thus, are gaining popularity for use in relay- and double-cropping systems. However, compared with summer biotypes, winter biotypes of camelina require a prolonged low temperature treatment (vernalization) to induce flowering. Research led to the discovery of a mutation in a gene coding for Flowering Locus C, a negative regulator of the flowering process, which is associated with the summer-annual biotypes of camelina. This discovery provides new knowledge pertaining to flowering/maturation time, which is an important trait for the development of double cropping systems. Thus, exploring additional regulators of flowering time could help identify ways to manipulate maturation time in oilseed cash crops and cover crops. This research also identified molecular markers for distinguishing summer- and winter-biotypes early in seedling development using a simple polymerase chain reaction technique to accurately classify biotypes of camelina prior to marketing. Additionally, we discovered significant differences in the freezing tolerance of summer- and winter-biotypes of camelina and conducted transcriptomic analyses to identify genes and signaling processes associated with differences in both cold-acclimation and freezing tolerance. Phenotypic analysis of offspring from a cross between a summer biotype (CO46) and a winter biotype (Joelle) suggest that as few as two genes may account for the bulk of the freezing tolerance in winter biotypes. Based on this data, an F7 Recombinant Inbred Line (RIL) population was developed from the offspring of this cross for map-based cloning of genes involved in these freezing tolerance and early flowering processes.
In collaboration with scientists at North Dakota State University (NDSU), we demonstrated that camelina planted in mid- to late-September had the best overwinter survival, spring stand establishment, and yield relative to earlier or later planting dates. Winter camelina planted into wheat stubble in Fargo, North Dakota, and Morris, Minnesota, in September of 2019 had excellent winter survival, with preliminary results from the growing season of 2020 showing greater than 90% weed suppression. For winter or summer camelina oilseeds being grown as a cash crop, determining the agronomic traits can be time consuming and expensive. In collaboration with scientists from NDSU, we developed methods for determining crude protein, total oil, and fatty acid profiles of camelina seed using near infrared spectrometry. These results will provide a rapid, non-destructive, high-throughput method for determining agronomically important traits in camelina seed from our recombinant inbred line population.
To meet the demand of the canola industry to increase production in the U.S., we also planted a bulk population of greater than 600 winter canola varieties into wheat stubble in Morris, Minnesota, and Fargo, North Dakota, on two planting dates (late August and mid-September) in 2019. More than seventy-five canola plants survived the winter at both locations and were genotyped and bagged for seed production. Because most plants that survived the winter were from the first planting date, the results indicate that plant development appears to be an important factor for freezing tolerance in canola. These results show promise for identifying winter hardy canola germplasm for increasing oilseed production in the U.S. or as an alternative option for multi-cropping systems.
Accomplishments
1. Development of a new diversity panel for winter canola genome wide association studies. Statistically associating genetic differences with how a plant looks or changes after a treatment is a powerful way to identify genes that control plant growth and stress responses. Once plant and genetic database resources are developed for such association studies, they can easily be used by others for many different research projects. ARS scientists in Fargo, North Dakota, successfully developed resources for a set of 429 varieties of winter canola, an important oil seed crop in the Midwestern U.S. and around the world. The resource was confirmed by identifying genes that control the speed of freezing tolerance in winter canola. Both the seedstock and genetic database from this set of plants have been shared with collaborators at North Dakota State University and Kansas State University, which provides canola breeders additional options to increase U.S. production of this important oilseed crop.
2. Identification of breeding tools associated with winter hardiness in canola. A lack of freezing tolerance in winter canola germplasm limits oilseed production in the upper Great Plains of the U.S. Identifying varieties with increased freezing tolerance and markers associated with this trait are needed by breeders to develop winter hardy canola varieties. ARS scientists in Fargo, North Dakota, characterized traits in a new breeding population of winter canola and used a genome wide association study approach to identify loci and markers associated with freezing tolerance. The thirteen genetic markers identified as being associated with freezing tolerance can be directly used to assist in genetic selection. The markers have been provided to breeders at North Dakota State University and Kansas State University, which will be used for improving freezing tolerance in elite lines of winter canola and to increase U.S. production of this important oilseed crop.
3. Approaches for creating weed tolerant crops. Weeds reduce the potential yield of corn just by growing near the crop plant - even before competition for resources such as light, nutrients, or water occurs. How a crop "sees" the weed, and how the crop reduces its growth in response is poorly understood. ARS scientists in Fargo, North Dakota, identified ten genes that were turned on or off when corn plants were grown with winter canola as a model for crop-weed interactions. This work also highlighted the probable role of salicylic acid in the corn response to weeds and led scientists to test the hypothesis that weed-induced salicylic acid promotes a crop response that alters the balance between growth and defense and, thus, causes yield loss. These genes will help scientists and industry find ways to engineer or breed corn to be blind to weeds, which will provide new options to customers and stakeholders for managing weeds and reducing yield losses due to competition from weeds.
Review Publications
Podder, S., Samarappuli, D., Anderson, J.V., Berti, M.T. 2020. Phenotyping a diverse collection of forage sorghum genotypes for chilling tolerance. Agronomy. 10(8):1074. https://doi.org/10.3390/agronomy10081074.
Bruggeman, S., Horvath, D.P., Fennell, A., Gonzalez-Hernandez, J., Clay, S.A. 2020. Teosinte (Zea mays ssp parviglumis) transcriptomic response to weed stress identifies similarities and differences between varieties and with modern corn varieties. PLoS One. 15(8):e0237715. https://doi.org/10.1371/journal.pone.0237715.
Arifuzzaman, M., Horvath, D.P., Rahman, M. 2020. Transcriptome analysis suggests cytokinin and gibberellin signaling may account for differences between spring and winter canola (B. napus) root development. Journal of Plant Biology. https://doi.org/10.1007/s12374-020-09270-6.
