Location: Plant Science Research
2022 Annual Report
Objectives
The overall goal of this project is to reduce nutrient inputs, particularly nitrogen (N) and phosphorus (P), in legume crops through the identification of germplasm having root architectural diversity and the discovery of genes that may contribute to that diversity. Desired outcomes from the research proposed herein include identification of unique germplasm with altered root morphology that may reduce costly fertilizer inputs, novel genes that regulate root development and function, and fundamental insight into the biochemical processes that affect nutrient acquisition. To achieve these goals and outcomes, three integrated objectives will be pursued.
Objective 1: Phenotype and evaluate root architecture changes in soybean, common bean and Medicago mutants, determine relationships between root architecture and improved nutrient acquisition, and define genome lesions.
Objective 2: Evaluate whole genome transcript analysis of common bean and alfalfa through RNA-seq analysis of roots, root nodules, leaves and seeds to compare wild-type and mutants.
Objective 3: Identify genes contributing to root architecture and nutrient acquisition in legumes and determine their function.
Approach
Identify mutant plants derived from fast neutron and Tnt1 mutagenized populations which affect root architecture and development, and define genetic lesions through next generation sequencing. Conduct RNA-seq transcript expression studies for the organs of wild type and mutant legume species such as alfalfa, common bean, and soybean to identify genes involved in unique adaptations displayed by these species. Utilize RNAi, zinc finger nuclease modification and/or antisense constructs to silence expression of selected root-specific/enhanced genes affecting root architecture and/or nutrient acquisition.
Progress Report
This is a bridging project that was initiated March 2018. Significant progress has been made in two objectives (1 and 3). In support of Objective 1, targeted mutagenesis was carried out on alfalfa to generate pho2 mutant plants that hyper-accumulate phosphate at rates of 3- to 6-fold higher than wildtype plants. A manuscript reporting these results was published. The next step is to test the hyper-accumulator plants in the field and two strategies have been initiated to further this goal. For the first strategy mutant plants were crossed with a high biomass line. Plants from this cross have been screened and clonally propagated to bulk seed for field plantings either this season or the next
In support of Objective 3, 600 root samples from 260 accessions of Medicago truncatula were phenotyped using laser ablation tomography (LAT), a method that allows for rapid high throughput analysis of the number and size of root cells and the composition of the cell walls using a specialized ultraviolet laser and camera system. The resulting data was processed and used in a genome-wide association study (GWAS) to identify candidate genes associated with variation in root-related traits. Work in the lab has focused on the construction of reagents to target these candidate genes as well as the optimization of the M. truncatula transformation protocol. In addition to these targets, candidate soybean root architecture related gene targets have been identified from published sources and reagent construction is currently in progress.
Significant progress has been made on the construction and validation of an enhanced gene editing reagent platform for legume plants. This platform encompasses both targeted mutagenesis and base editing strategies. Several candidate gene targets in soybean, alfalfa, and M. truncatula have been identified and their reagents generated and tested in transient transformation assays.
Accomplishments
1. The SELF PRUNING 3C gene controls flowering time and root growth in tomato. The genetic basis of many agronomically important traits involve variation in members of the CENTRORADIALIS, TERMINAL FLOWER 1, SELF PRUNING (CETS) gene family. ARS researchers at St. Paul, Minnesota, and collaborators at the Universidade Federal de Viçosa and Universidade de Sa~o Paulo in Brazil found that one member of this family, the SP3C gene, acts as a flowering repressor and modulates root growth. In mutant plants where spc3 gene function has been disabled, accelerated seed germination, and increased root length with reduced lateral branching was observed, while when SP3C is over-expressed in transgenic lines the opposite effect was observed. These discoveries provide new insights into the role of this gene in root-related traits and support future exploration of the role of this gene family in other crops including legumes for plant breeders to develop plant cultivars that establish more rapidly due to rapid seed germination and have greater drought tolerance from increased root length.
Review Publications
Del Mar Martinez-Prada, M., Curtin, S.J., Gutierrez-Gonzalez, J.J. 2022. Potato improvement through genetic engineering. GM Crops & Food. 12(1):479-496. https://doi.org/10.1080/21645698.2021.1993688.
Miller, S.M., Dornbusch, M.R., Farmer, A., Huertas, R., Gutierrez-Gonzalez, J.J., Young, N.D., Samac, D.A., Curtin, S.J. 2022. Alfalfa (Medicago sativa L.) pho2 mutant plants hyperaccumulate phosphate. G3, Genes/Genomes/Genetics. 12(6). Article jkac096. https://doi.org/10.1093/g3journal/jkac096.
Curtin, S.J., Qi, Y., Peres, L., Fernie, A.R., Zsogon, A. 2021. Pathways to de novo domestication of crop wild relatives. Plant Physiology. 188(4):1746-1756. https://doi.org/10.1093/plphys/kiab554.
