ASHS 2024 Conference

In this interactive workshop, participants will be provided training on how to use Rosaceae (GDR, www.rosaceae.org), Vaccinium (GDV, www.vaccinoum.org) and Citrus (www.citrusgenomedb.org) specialty crop databases resources, as well as the Breeding Data Management tools, BIMS (www.breedwithbims.org), and the Field Book App for phenotype data collection. Using a case-study approach we will focus on how to use these integrated resources most efficiently for research and crop improvement efforts, how to apply FAIR data principles to sharing and submitting research data to these databases at the time of publication and facilitate a robust dialogue between researcher, breeders and the development team on needed improvements and long-term sustainability options for these resources.

This interactive workshop will bring together researchers and breeders to accelerate research and crop improvement in specialty crops. It will provide training and feedback on how to most effectively use the NIFA SCRI and National Research Support project 10 (NRSP10) funded Rosaceae, Citrus, and Vaccinium specialty crop databases and breeding data management resources (BIMS and Field Book), learn how to follow FAIR data principles, foster better two-way communication for increased feedback from researchers on further development of these resources, and continue to build a robust community around these research and crop improvement tools. The trainers include users and developers of these resources, ranging from early career to well-known and respected research enabling resource generators.

The Genome Database for Rosaceae (GDR), Genome Database for Vaccinium, and Citrus Genome Database (CGD) offer critical resources and tools to enable genomic, genetic, and breeding research for fruit, nut, and ornamental crops of great economic importance to the U.S. In addition, BIMS (BreedwithBIMS.org) and the Field Book APP provide widely used tools for managing plant breeding program data. While these resources are heavily used worldwide, many researchers are unaware of the full potential of using them and how they can contribute their own data for wider recognition and reuse. This interactive workshop aims to bridge this knowledge gap by providing hands-on training for specialty crop researchers on how to best use these resources and contribute their data. The workshop will also solicit ideas from participants on how to improve these databases and discuss the various options to make them sustainable in the long term. The workshop will bolster the utilization of integrated big data, promote future data sharing, and ensure that data is FAIR (Findable, Accessible, Interoperable, and Reusable).

The workshop aims to bring together researchers to accelerate research by more effective use of specialty crop databases and breeding data management resources, encouraging data submission at the time of publication, and gaining feedback from researchers. Through hands-on training, participants will become more familiar with the database resources and breeding data management tools, learn how to submit their data, and provide essential input for improving these databases and their long-term sustainability.

Coordinator(s)

• Dorrie Main, Washington State University, Pullman, WA, United States

Moderator(s)

• Cameron Peace, Washington State University, Horticulture, Pullman, WA, United States

Speaker/Participant(s)

• Dorrie Main, Washington State University, Pullman, WA, United States

Introduction to the Workshop and Specialty Crop Research Databases (15 mins)

• Jodi Humann, Washington State University, Horticulture, Pullman, WA, United States

How to efficiently use integrated genomics data and tools for research (20 mins)

• Sook Jung, Washington State University, Horticulture, Pullman, WA, United States
  How to efficiently use integrated genetics data and tools for research (20 mins)
  
  
• Nahla Bassil, USDA ARS, National Clonal Germplasm Repository, Corvallis, OR, United States
  How to efficiently use integrated germplasm and genotype data for research (15 mins)
  Summary:
  
  
• Trevor Rife, Clemson University, Plant and Environmental Sciences Department, Florence, SC, United States
  How to use the Field Book App for phenotypic data collection (15 mins)
• Sushan Ru, Auburn University, Auburn, United States
  How to use the Breeding Information Management System, BIMS, for Crop Improvement (20 mins)
• Cameron Peace, Washington State University, Horticulture, Pullman, WA, United States

Revolutionizing Grapevine Breeding: Overcoming Perennial Challenges with Genome Editing - Bridget Bolt
CRISPR/Cas9-Mediated Development of Low Mowing Frequency Perennial Ryegrass and Tall Fescue Varieties - Roshani Budhathoki
Precision Breeding Using CRISPR to Improve Production Traits in Blackberry - Pradeep Marri
Double CRISPR Knockout of Pectin Degrading Enzymes Improves Tomato Shelf-life While Ensuring Fruit Quality - Isabel Ortega Salazar
Functional Characterization of a Candidate Bacterial Wilt Resistance Gene in Tomato - James Duduit
Novel Short Synthetic Promoters for Constitutive Expression in Dicot Species - debao huang

Climate change poses a significant threat to perennial crops like grapevine in the USA, with projections indicating that 50-81% of acreage may become unproductive by 2040. To address this, breeding resilient varieties is imperative. However, traditional breeding for perennials is time-consuming (25-30 years) due to extended evaluation periods. Genome Editing, specifically CRISPR/Cas9, offers targeted modification potential but faces hurdles in perennial crops such as grapevine. These include somaclonal variation, inefficient transformation, transgene removal needs, and industry hesitance. We propose an efficient Genome Editing method for grapevine leveraging the systemic mobility of methylated dicistronic mRNA:tRNA molecules. This approach aims to: 1. Enable genome editing without transgene integration, negating the need for backcrossing and preventing unintended genetic changes. 2. Circumvent in-vitro culture, reducing somaclonal variation risk and preserving existing chimerism. Our method addresses key challenges in Genome Editing adoption for grapevine and could accelerate the development of climate-resilient varieties, crucial for sustainable agriculture in a changing climate.

Tall fescue (Lolium arundinacea Schreb.) and perennial ryegrass (Lolium perenne) are common cool-season turfgrass species. They are widely utilized in home lawns, athletic fields, golf courses, and roadsides. However, these grasses require frequent mowing to maintain lawn quality. Developing low-mowing frequency varieties is highly desirable because it can significantly reduce maintenance costs and fuel consumption. Gibberellin 20-oxidases (GA20ox) are key genes in the gibberellic acid (GA) biosynthesis pathway, mutations in which can induce a dwarf phenotype in plants and, therefore, reduce mowing frequency, as demonstrated by our dwarf turf varieties developed using conventional mutagenesis techniques. We have recently employed CRISPR/Cas9 technology to create knockout or knockdown mutations in the GA20oxidase1 gene to develop dwarf tall fescue and perennial ryegrass lines. The CRISPR/Cas9 construct used for editing LaGA20ox1 and LpGA20ox1 has ZmUbi and OsU6a promoters controlling the expression of Cas9 and sgRNA, respectively. We have delivered the CRISPR/Cas9 construct into tall fescue and perennial ryegrass via Agrobacterium-mediated transformation. We have observed a 35-50% reduction in plant height compared to the parental wild-type plants. Furthermore, the leaf widths in these mutants are reduced by 30-60%. Some mutants display a smoother leaf texture compared to the wild type. The T2 homozygous mutant progenies will undergo field evaluation for mowing frequency and fertilizer requirements. We anticipate that some of these gene-edited lines should exhibit a significant reduction in mowing frequency and fertilizer input.

Pairwise is a technology-focused food and agriculture company that makes tiny alterations to plant genetics to solve big problems. Powered by our best-in-class technology, Pairwise is harnessing new genomics technologies to create innovative new products. With our FulcrumTM platform, we accelerate innovation in plants and plant-based production systems, delivering value to people and our planet. Partnering with Plant Sciences, Inc. to access their elite germplasm, we created a platform to transform and edit tetraploid blackberry (Rubus sub. Rubus). We used the power of genetics and genomic tools to identify the candidate gene for thornless in blackberries and used CRISPR based gene editing to validate the trait. The identification of this gene will enable faster deployment of a thornless trait, an essential production trait, into blackberries. The gene may be broadly applicable to other Rubus species and horticultural crops such as roses.

Tomato fruit is an important and popular commodity worldwide. One of the main challenges for the fresh market tomato industry is postharvest deterioration, which is mainly determined by the rate of softening. This rate can affect tomato shelf-life, pathogen susceptibility, and fruit waste. The softening rate is regulated by multiple factors, but mainly by the pectin composition of the cell wall, which is remodeled, disassembled, and solubilized by enzymes during fruit ripening; a process that induces and determines the fruit softening in fleshy fruits. The main pectin-degrading enzymes that act on the pectin backbone are Polygalacturonases (PG) and pectate lyases (PL). In this study, we generated a double CRISPR knockout PGPL. We investigated the combined functions of SlPG2a and SlPL on fruit quality traits in postharvest, including shelf-life attributes like firmness and water loss, fruit marketability, and disease incidence. We also assessed additional attributes impacting consumer acceptance, such as taste and aroma. Our findings revealed that the tomato ripening enzymes SlPG2a and SlPL act additively, significantly affecting fruit firmness and shelf-life. Additionally, aspects of fruit quality, such as external color, sugar: acid ratio, and aroma volatiles, were improved or not affected in the double CRISPR knockout PGPL when compared to control. The discoveries of this research provide new insights into the influence of pectin backbone degradation on fruit physiology and postharvest quality, which can be used in crop improvement programs to make fruit more resilient in the supply chain without compromising consumer-based quality traits.

Bacterial wilt, caused by Ralstonia solanacearum (Rs), is an economically devastating plant pathogen that causes rapid death and has been widely distributed worldwide. Rs is a soil-borne bacterium that plugs plant xylem vessels, causing wilt and ultimately death in tomatoes (Solanum lycopersicum L.) and many other economically important crops. The most cost-effective and efficient means of managing Rs is planting resistant cultivars. However, acceptable Rs resistance in these genotypes is tightly linked to small fruit size, preventing development of BW-resistant large fruited tomato cultivars. Previous research has shown that a highly resistant cultivar's candidate resistance gene (Sl-BWR) is linked to qualitative resistance of Rs. Our preliminary data indicates that overexpression of the resistant allele in a susceptible cultivar background confers resistance comparable to the wild-type resistant allele. There is ongoing work to generate susceptible allele overexpression lines and knockout lines from the susceptible and resistant cultivars. We hypothesize the resistant allele knockouts will be susceptible to Rs, indicating that the candidate gene is the primary resistance factor. The gene could be functionally characterized to elucidate the Rs resistance mechanism in tomatoes to be deployed in a breeding program to develop resistant cultivars against bacterial wilt.

Determining the promoter motifs involved in regulating transcription, the first process of gene expression, is critical for synthetic promoter engineering. A gene’s promoter contains cis-regulatory elements, or motifs, which are binding sites for transcription factors (TFs) to initiate and drive transcription. Many bioinformatic tools have been developed for determining statistically overrepresented regions, representing sites of potential cis-regulatory elements, that are shared across groups of promoters. Combining the results produced by multiple bioinformatic tools can lead to improved detection accuracy of motifs conferring biological activity. In the present study, we compiled a set of 11 known soybean constitutive gene promoters under the assumption that some of the promoters are regulated by the same transcription factor(s). Seven bioinformatic tools capable of de novo motif discovery were used to determine potentially shared motifs within the promoters, which were then mapped back to the original promoter sequences. A total of 64 overlapping motif regions (OMRs) were commonly detected amongst the 11 constitutive promoters, and each OMR was cloned individually in front of the minimal CaMV 35S promoter driving GUSPlus reporter gene expression. Transient tobacco leaf agroinfiltration and subsequent quantitative GUS activity assays were used to determine each OMR’s ability to drive reporter gene expression. We found that 20 of the 64 bioinformatically-determined OMRs drove functional gene expression significantly higher than the basal levels conferred by the minimal 35S promoter. Of the 20 functional OMRs, 11 drove GUSPlus expression at levels from just twice that of the minimal 35S promoter up to nearly half of the full-length 35S promoter. We also transformed these 20 functional OMRs individually into Arabidopsis. While GUS staining is still ongoing, we identified a few OMRs which showed strong promoter strength in single-copied homozygous Arabidopsis seedlings. These functional OMRs are strong candidates for further characterization and can be used for crop improvement.

Sponsoring Professional Interest Groups
Technology: Coordinator Milt McGiffen - milt.mcgiffen@ucr.edu
Teaching Methods: Coordinator, Kathryn Orvis – orvis@purdue.edu
Controlled Environment: Coordinator, Kent Kobayashi - kentko@hawaii.edu

Supporting Professional Interest Groups
Federal Partners: Matthew Mattia - Matthew.Mattia@usda.gov
Plant Biotech: Kedong Da - kda@ncsu.edu
Ornamentals/Landscape and Turf; Youping Sun - youping.sun@usu.edu
Local Food Systems: Charles H. Parrish II - chip.parrish@pm.me

Artificial intelligence and related topics, e.g., robotics, have been a long time coming in agriculture. For decades there have been predictions of intelligent robots replacing humans, and large farms run by a few humans with many autonomous tractors and other devices. But with the now widespread use of artificial intelligence in everyday life,
the moment has arrived. We developed this colloquium by casting a wide net out to all the Professional Interest Group Chairs, and have assembled talks and demonstrations from general topics to specific applications.

Two online meetings were held, where Professional Interest Groups officers and those interested suggested speakers and discussed topics. Further discussions over email helped fill in the details to create this colloquium.

We will have a block of speakers for the diverse topics we present below, as well as panel discussions on how AI is and can be incorporated into various aspects of Horticulture, so that there is ample time for questions and discussion.

Title: Overview of the Colloquium

Speaker: Milt McGiffen, Cooperative Extension Specialist, Department of Botany and Plant Sciences,
University of California, Riverside, CA.

AI in Ornamentals

Title: FloraCount: An App for Rapid Assessment of Pollinator Attractiveness to Annuals and Perennial Plants.

Description: Customers are interested in buying annuals and perennials that support pollinators. Protocols for rapid assessment in flower trail evaluations are not available. We have developed a mobile app that can be used to analyze in real time the users’ observational data and quantitatively rank the relative utility of observed cultivars to pollinator communities. This app takes into account pollinator groups, relevant floral characteristics and landscape.

Presenter: Harland Patch
Assistant Research Professor
Department of Entomology
Penn State University
549 Ag Sciences & Industries Building
University Park, PA 16802

Title: Approach to Biodiversity Protection: Employing AI and IoT Systems for the
Containment of Box Tree Moth Proliferation.

Description: The box tree moth (BTM, Cydalima perspectalis) is an invasive pest first confirmed in Niagara County, New York in 2021. This invasive pest can significantly damage and potentially kill boxwood (Buxus species) plants if left unchecked. This presentation describes our advances in combining deep learning algorithms for enhanced computer vision with IoT-enabled smart traps, to facilitate the early detection and continuous monitoring of BTM populations and to protect the prevalent ornamental boxwood in U.S. landscapes.

Presenter: Yanqiu Yang (she/her)
Ph.D. Graduate Research Assistant
Department of Agricultural and Biological Engineering
Pennsylvania State University
3 Agricultural Engineering Building
University Park, PA 16802

Title: Landscapes from Words: The Future of Landscape Design with AI.

Description: The ongoing text-to-graphic artificial intelligence (AI) revolution has the potential to change the field of Landscape Architecture dramatically. The ability to produce original high-quality graphics, manipulate the viewer's perspective of images, and amend the rendering style through text inputs are significant advancements that will
inform new design process models. These changes can lead to expanded design exploration, improved accessibility for non-designers to contribute to creating visual concepts, enhanced ability to integrate data analysis and visualizations, and streamlined collaboration between clients and project stakeholders using a shared visual language. This talk focuses on two dimensions of change that may result from the rapid evolution of text-to-graphic AI, including (1) faster iterations and exploration of design options and (2) the advancement of methods that result in more inclusive and responsive design. In the classroom, students are just beginning to acknowledge the existence of text-to-graphic AI, which allows them to experiment with text-based design options that allow them to quickly visualize and explore a wide range of site program alternatives. Nevertheless, how do we manage the ethical and creative boundaries within an academic setting? In a research context, methods supporting rapid manipulation of both generated images and existing landscape photography represent advances that allow for greater collaboration surrounding landscape design decisions (Incorporating resilience strategies, protecting vernacular landscape elements that support a sense of place, or representing new design proposals that modify the landscape). These approaches allow stakeholders to gain remarkable advances in influencing the design process through shared visualization development. However, as with any emerging technology, practitioners, educators, and researchers need to respond to the challenges presented by text-to-graphic AI by developing and testing new design process models and public engagement techniques that can improve landscape decision-making and streamline collaboration.

Presenter: Aaron Thompson
Assistant Professor
Department of Horticulture and Landscape Architecture
Purdue University
625 Ag Mall Drive
West Lafayette, IN 47906

Title: Developing Guidelines for Extension’s Use of ChatGPT and Other Generative AI
Tools.

Description: A new technological era marked by the advent of Artificial Intelligence (AI), particularly generative AI and Large Language Models (LLMs) like ChatGPT has necessitated the need to navigate this domain with a compass of ethicality, safety, and effectiveness. Penn State’s experience developing guidelines for Extension’s use of
generative AI tools which will be shared and discussed.

Presenter: Michael Masiuk
Assistant Director – Horticulture Programs
Penn State Extension
342 Agricultural Administration Building
University Park, PA 16802

Panel: 30 minute panel with the above speakers, to allow time for Q&A and discussion.