Progress 08/01/24 to 07/31/25

Outputs
Target Audience:Our project engaged a varied set of stakeholders during this reporting period, spanning undergraduate education and the professional research community. Undergraduate Training: We provided hands-on research training to two undergraduate students from Oregon State University's Botany program: Piper Kelton and Luc Stone. Oregon State University is a state university which supports agricultural education, training, and research for the state of Oregon and Pacific Northwest.These students were actively involved in experimental design, data collection, and analysis. Their participation fostered early-career development in plant biology and strengthened the pipeline of future agricultural researchers. Professional Research Community: Project results and methodologies were shared with the broader scientific community through participation in the Maize Genetics Conference, a leading venue for advances in plant genetics and breeding. The Maize Genetics Conference is organized by the Maize Genetics Cooperation, a professional society and network of agricultural researchers focused on understanding and improving our nation's most important crop. Engagement at this conference facilitated peer feedback, collaborative opportunities, and dissemination of findings to researchers focused on crop improvement and translational plant science. These outreach efforts support both workforce development and knowledge transfer in line with USDA's goals to strengthen U.S. agriculture through education and innovation. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Involved research staff have had opportunities for both training and professional development. Postdoctoral Scholar, Brian Zebosi learned advanced bioinformatics and genomics through the conducted work. He advanced his professional development by presenting a research poster at the Maize Genetics Conference. During this project he has mentored and trained two undergraduate researchers, which is strongly aligned with his objectives of becoming a tenure-track professor at an agricultural research institute. Undergraduate researchers Piper Kelton and Luc Stone have recieved hands-on training in agricultural sciences, including field, greenhouse, and lab work with genetic mutants and molecular biology. Kelton presented her work at an undergraduate research symposium as a poster, advancing her professional development in the agricultural sciences. How have the results been disseminated to communities of interest?Project results have been disseminated to two primary communities of interest: (1) the professional research community focused on maize genetics and (2) undergraduate students engaged in agricultural research training. These outreach activities support both the advancement of scientific knowledge and the development of a skilled agricultural workforce. Dissemination to the Scientific Community: Scientific findings from the project were shared through a research poster presented at the Maize Genetics Conference, an annual international meeting that brings together academic researchers, industry scientists, and plant breeders working on fundamental and applied questions in maize biology. The poster, presented by project participant Brian Zebosi, communicated early-stage discoveries related to carpel suppression gene mapping and characterization in maize. Sharing these findings in a collaborative research setting enabled peer feedback, fostered potential partnerships, and accelerated the integration of our discoveries into the broader maize research community. Undergraduate Research Training and Outreach: We also disseminated results through hands-on undergraduate research experiences, an important outreach channel for expanding access to agricultural science and building future research capacity. Two Oregon State University botany students, Piper Kelton and Luc Stone, were trained in experimental design, genetic screening, and phenotypic analysis. As part of her research experience, Piper Kelton developed and presented a scientific poster summarizing her findings, contributing to her academic growth and professional development. These training experiences help demystify research for undergraduate students and increase retention in science, technology, and agricultural career pathways. By integrating formal scientific communication with undergraduate mentoring and training, our project has contributed to the dual goals of advancing agricultural research and fostering public engagement in plant science. Future outreach efforts will continue to emphasize training opportunities and engagement with both academic and broader agricultural communities. What do you plan to do during the next reporting period to accomplish the goals?In the next reporting period, our focus will be on advancing all three project objectives through continued genetic mapping, molecular characterization, transcriptomic analysis, and comparative phenotyping. These efforts will deepen our understanding of carpel suppression and grain production in maize and its relatives, while addressing key questions that remain from the current reporting period. We will prioritize clear, testable hypotheses, increase experimental efficiency by refining our allele selection, and generate high-impact datasets to support upcoming publications and long-term crop improvement goals. To support our goal of characterizing 40 carpel suppression mutations, we will continue to use our mapping pipeline to identify new alleles. Although we have been successful in our mapping efforts so far, we will omit alleles with low penetrance or inconsistent inheritance to maximise our chances of success during mapping. With the alleles in hand, we will continue to characterize potential candidate genes, quickly validating potential new alleles of tasselseed1 and tasselseed2 by allele complementation tests, hormone treatments, and Sanger sequencing of gene coding regions. These data are in preparation for a peer reviewed publication to be submitted in late 2025. In the next review period we will perform RNA sequencing on theTasselseed3 mutant and wild-type plants and use transcriptomic analysis techniques to position Tasselseed3 within known genetic floral development pathways. We will treat Tasselseed3 plants with synthetic plant hormones and perform hormone level measurements to validate potential mechanisms by which Tasselseed3 impacts carpel suppression and plant fertility. We will analyze X-ray CT data collected for carpel suppression mutants created in Setaria and compare the quantitative phenotype with maize to draw conclusions about the conservation of grain productionpathways and open up avenues for their improvement across cereal crops. Together, these activities will build a strong foundation for defining the molecular basis of carpel suppression, identifying conserved mechanisms across crop species, and developing potential genome editing targets for yield improvement. In addition, these research efforts will provide valuable training opportunities for undergraduate and postdoctoral researchers involved in the project, ensuring continuity of skills development and scientific workforce training.

Impacts
What was accomplished under these goals? Grain crops like maize and millets are essential to global food security, yet their ability to produce grain is limited by a natural process called carpel suppression. In grasses like maize, about half of all flowers fail to produce grain due to this suppression. By understanding this process, we aim to improve crop yield potential through targeted genome editing. This project focuses on identifying the genes that control carpel suppression in maize and related grasses and understanding how these genes work together. Over the past year, we made substantial progress toward this goal through three major objectives: 1) mapping carpel suppression genes in maize, 2) uncovering how these genes interact in regulatory pathways, and 3) testing whether similar genes affect grain production in related crops like Setaria, a model for millets. In this reporting period, we significantly advanced objectives 1-2 and have positioned ourselves to achieve objective 3 in the next funding period. We furthered our objective of understanding the genes and genetic pathways that control grain production by conducting gene mapping and experimental genetics in maize. In 2024-2025, we characterized 14 genetic mutants of maize with abnormal carpel suppression. We constructed maize populations segregating these mutations and isolated DNA from pools of mutant and wild-type plants. Using high throughput DNA sequencing technology, we produced 7.7 billion sequencing reads representing 1.161 trillion nucleotides from the maize genome. We identified genetic mapping intervals for each of the 14 new mutant alleles. Our sequencing data suggests that 3 are novel alleles of the classic mutant tasselseed1 and 3 are novel alleles of tasselseed2, but the remaining 8 are linked to genes which have not been previously implicated in crop flower development. To conduct this genetic mapping research, Postdoctoral Scientist, Dr. Brian Zebosi, developed a new gene mapping pipeline. His software and scripting tools are currently in preparation as a peer reviewed publication and will be released, open access to the benefit of the crop research community and public. We furthered our objective of determining interactions between carpel suppression pathways in maize through the detailed characterization of the classic mutant Tasselseed3, for which the causal gene and mutation is currently unknown. Using our mapping interval data and dense sequencing information, we developed a series of 12 molecular markers which we used to identify mutant individuals prior to the manifestation of mutant phenotypes. Work was led by Piper Kelton, an undergraduate researcher, advancing our goal of agricultural workforce development. We used this genotyping assay to set up an RNA sequencing experiment which we will use to understand the RNA transcripts which precede the Tasselseed3 phenotype and will help determine the molecular mechanism of this poorly understood mutant variety via transcriptomic analysis. We are also using our molecular marker to prepare hormone treatment experiments with jasmonic acid (JA) and brassinolide (BL) to understand the interaction between the Tasselseed3 phenotype and hormone pathways that are already implicated in flower development. To further our work on objective 3 we have recently searched for and put staff in place to conduct the proposed X-ray CT analysis in PD Leiboff and collaborator Bartlett's research groups. CT data have already been collected and so staffing this postdoctoral level position with a research scientist who is already an expert in X-ray CT data analysis will allow us to complete our research objectives within the next review period. Our work this year has significantly expanded the scientific foundation needed to address carpel suppression in cereal crops. We identified promising new alleles that control this trait in maize, laid the groundwork for uncovering how they interact in genetic pathways, and have positioned ourselves to translate this knowledge to other important crops through genome editing. These discoveries not only provide tools for researchers and breeders to improve yields, but also contribute to training the next generation of agricultural scientists. Undergraduate and postdoctoral researchers were actively involved in every stage of this research, gaining skills in genetics, genomics, and plant biology. By advancing both knowledge and workforce development, our project supports long-term agricultural resilience and yield improvement.

Publications