Location: Plant Science Research
2021 Annual Report
Objectives
Crop improvement is a balancing act requiring simultaneous selection for multiple diverse traits, including resistance to a range of diseases, to develop superior new cultivars. One of the diseases that is a subject of investigation here (Fusarium head blight) continues to cause significant economic losses to the U.S. wheat crop, while another (stem rust) has the potential to do so. Similarly, crown rust continues to be a significant disease of oat. The overall goal of this project is to use genetic engineering technologies to develop novel molecular variants of specific genes and validate that they, as well as a previously identified spontaneous mutation, improve resistance to these particular diseases in wheat and oat. The approaches for improving disease resistance will generate novel resources and knowledge for protecting wheat against both FHB and stem rust, and oat against crown rust, in a manner that complements current breeding efforts for both diseases. These research activities will be coupled with the coordination of a service activity that provides a conduit for Midwestern hard red spring wheat breeders to evaluate jointly their advanced germplasm for agronomic quality and disease resistance at multiple locations. Combining basic and applied research in this manner will ensure that new wheat and oat cultivars retain high yield and quality while also being protected from current and potential disease threats. To achieve project goals, three objectives will be pursued:
Objective 1: Evaluate a novel wheat genome deletion that improves Fusarium head blight resistance in adapted hard red spring wheat under field conditions.
Sub-Objective 1.A. Evaluate the effect of genetic background on Fusarium head blight resistance conferred by a novel genome deletion.
Sub-Objective 1.B. Evaluate the effect of pyramiding the deletion and the partial FHB resistance gene Fhb1 on suppression of FHB.
Sub-Objective 1.C. Evaluate the effect of the deletion on agronomic performance in contemporary hard red spring wheat.
Objective 2: Establish efficient transformation systems in parallel for wheat and oats, and improve disease resistance by endogenous gene disruption and foreign gene addition.
Sub-Objective 2.A. Validate candidate rust susceptibility genes in the model grass Brachypodium.
Sub-Objective 2.B. Disrupt stem rust susceptibility genes in wheat.
Sub-Objective 2.C. Disrupt crown rust susceptibility genes in oat.
Objective 3: Coordinate the Hard Red Spring Wheat Uniform Regional Performance Nursery Program.
Approach
Objective 1 seeks to enhance Fusarium head blight resistance in hard red spring wheat by introducing a unique genome deletion that improves resistance to this disease. We will determine if the deletion will improve resistance in other susceptible wheat genotypes. Near-isogenic lines of two susceptible hard red spring wheat cultivars that either possess or do not possess the deletion have been developed. These lines will be evaluated in Fusarium head blight nurseries at several locations to determine if lines with the deletion exhibit improved Fusarium head blight resistance when compared to the lines that do not possess it. We will test whether the deletion, when paired with a Fusarium head blight resistance gene, enhances Fusarium head blight resistance synergistically. Near-isogenic lines of two Fusarium head blight-susceptible hard red spring wheat cultivars that possess either the resistance gene alone or the gene together with the deletion will be evaluated in Fusarium head blight nurseries to determine if lines with both the deletion and the resistance gene exhibit superior resistance compared to the lines with resistance agene alone. We will examine how the deletion affects agronomic performance. The deletion has been introduced into diverse hard red spring wheat breeding lines. These near-isogenic lines and the original parents will be grown in field plots at several locations. Agronomic traits will be measured in the near-isogenic lines and compared to their parents to determine if the deletion has a detrimental effect on them. Objective 2 seeks to improve resistance to wheat stem rust and oat crown rust. We will employ the model grass Brachypodium as a testbed to test if mutating certain genes enhances resistance to these diseases. Genome editing using the CRISPR/Cas9 system will be used to perturb the genes, which are known or thought to enhance resistance to pathogens in other plant species when disrupted, in Brachypodium. Plants with confirmed mutations in the genes will be inoculated with the pathogens that cause wheat stem rust and oat crown rust, to confirm that their disruption improves resistance to these diseases. We will build on these results by creating, in wheat, mutations in the genes that enhance stem rust resistance in Brachypodium, and determining whether they also improve stem rust resistance in wheat. We will also create mutations in these same genes but in oats, to determine whether enhanced crown rust resistance can be obtained. Objective 3 will provide hard red spring wheat breeding programs in the upper Midwest an annual opportunity to have their advanced wheat germplasm evaluated for performance at more than a dozen field sites in fives states and Canada. The advanced lines are planted in replicated plots at these locations, and agronomic trait data on the germplasm are obtained by colleagues at each location.
Progress Report
In FY2021, progress was made on several research projects that comprise the project plan. For Objective 1, wheat near-isogenic lines that carry a chromosomal deletion that enhances resistance to the fungal disease Fusarium head blight in greenhouse experiments were evaluated for resistance in field disease nurseries located in three different states in the Upper Midwest. In 2020, the second year of these evaluations was completed. A cursory statistical analysis of the data suggests that under the high disease pressure of these field disease nurseries, the deletion does not significantly improve Fusarium head blight resistance. Similarly, combining this deletion with the wheat Fhb1 gene that confers partial resistance to Fusarium head blight resistance does not significantly enhance resistance beyond that contributed by Fhb1 alone under field conditions.
A second project within Objective 1 seeks to determine how the aforementioned deletion impacts agronomic properties of wheat. The deletion has been introduced into several elite wheat genotypes from spring wheat breeding programs at three Midwestern universities. Grain of these genotypes was planted in replicated field plots by university collaborators at locations in North Dakota and South Dakota, and agronomic properties will be evaluated on these plots to determine if any may be negatively impacted by the presence of the deletion. Statistical analysis of data on a related project with an ARS colleague at Fargo, North Dakota was completed and determined that milling and baking properties are not altered due to the deletion, while also revealing that the deletion has a positive effect by increasing flour protein content significantly.
A different area of research being pursued as part of the project plan involves the use of gene editing to improve disease resistance in wheat and oats, by disrupting so-called disease susceptibility genes. This research represents Objective 2. Proof of concept studies with the model grass Brachypodium distachyon have advanced, with each of the three target genes selected for the project having successfully been disrupted by a well-established gene editing method. For FY2021, a second generation of these lines was obtained by selfing, and these were genotyped to identify plants that no longer have the T-DNA insert, as well as to identify plants that are homozygous for particular editing events that disrupt the target gene. Plants with gene editing events in one of each of the three genes selected for disruption are currently being evaluated for their effect on the development of stem rust disease symptoms, in an effort to determine if they are candidates for gene editing in wheat and oats to improve resistance to stem rust.
A second area of research under Objective 2 is to develop gene editing tools for wheat and oats. Research to develop an efficient transformation method for oats is ongoing. This will be critical for proposed gene editing research in this crop species. A new method for Agrobacterium-mediated transformation, namely the use of young oat stem segments as the tissue of choice for transformation, was pursued. This method is far simpler and faster than the two methods that were being pursued previously, and thus should be more efficient as well. To date, no transformants have been recovered, and so new plasmid vectors are being developed with the hope that this will result in the recovery of transformed plants and thus establishment of a key method with broad relevance to oat improvement.
Lastly, Objective 3 addresses annual coordination activities associated with the Hard Red Spring Wheat Uniform Regional Performance Nursery program. The final step in the coordination of the 2020 nursery, involving the compilation and analysis of agronomic performance data of elite wheat germplasm at multiple locations in four states, and development and dissemination of a final report, was completed. The initial stages of coordinating this nursery program for 2021, which involved solicitation of entries from prospective participants and then orchestrating seed exchange of these entries among location cooperators, was completed. Similarly, the annual report of a second nursery program, the 2020 Uniform Regional Scab Nursery, was completed and distributed to participants, and the 2021 year of this nursery program was coordinated in the same manner described above.
Accomplishments
1. A mutation improves the protein content of bread wheat. Wheat is a key source of protein for humans globally but yield and quality of grain are impacted by diseases such as Fusarium head blight. ARS scientists in Saint Paul, Minnesota, and Fargo, North Dakota, evaluated wheat grain from plants with a mutation that improves resistance to Fusarium head blight to determine if the mutation has an effect on grain quality or end use properties. The mutation does not alter grain and kernel physical characteristics, milling properties, or functional properties of the flour in baking but significantly increases protein levels in the grain and flour. These results suggest that the mutation may eliminate a gene that negatively regulates seed protein accumulation. Wheat lines with the mutation are valuable resources for breeding wheat with enhanced resistance to Fusarium head blight and can add value to the grain by increasing protein content.
