Assembling a Reference Panel of DNA Profiles for U.S. Heirloom Apple Cultivars - Cameron Peace Rediscovering Lost Heirloom Apple Cultivars with DNA Fingerprinting - Dongyun Lee Air or Soil Temperature: Understanding the Cues for Dormancy Transition in Peach - Ksenija Gasic Selection and Evaluation of Citrus Resistobiome for HLB Resistance/Tolerance - Yongping Duan Population Genetics and Genome-wide Association Studies Provide Insights into the Genetic Basis of Persea Fruit Quality Traits - Gul Ali Genome Assembly of Persia Americana cv. Simmonds Provides Insights on Genetic Relationships Among Avocado Hybrids Exhibiting Tolerance To Laurel Wilt - Vincent Njung'e Michael Utilizing Haploid Pollen Grains and Diploid Leaf Tissue Genomic Sequence Data to Phase the ‘Wonderful’ Pomegranate Genome - Giuseppe Lana
Heirloom apple cultivars represent an important specialty crop for producers and a genetic resource for the dessert apple industry. These cultivars are plagued with misidentifications, which hinders utilization and long-term preservation. Phenotypic identification, used for centuries, is unable to distinguish among the thousands of existing U.S. heirlooms. DNA profiling provides an objective basis for cultivar identification. Washington State University’s “MyFruitTree” (myfruittree.org), built upon the RosBREED project and with international collaborations, has accumulated a DNA profile dataset of thousands of apple individuals focused on the U.S. genepool. Users submit leaf samples for trees of interest, and MyFruitTree’s cost-recovery research opportunity determines the cultivar identity (or reveals their uniqueness and pedigree position). However, a common and valid question is, “How do you know that is the correct identity?” The core panel of robustly identified cultivars was based on public breeding germplasm, modern cultivars, and their ancestors, which were DNA profiled in the RosBREED project last decade. Since then, examined trees from collections in the U.S. and abroad have greatly expanded the number of DNA profiles with cultivar labels. But those labels are not always correct, especially when a tree is derived from only a single source. Therefore, a system was derived for assigning confidence to the cultivar labels associated with DNA profiles. Cultivar name evidence is assembled in the categories of provenance, phenotype, and genotype for both the DNA-profiled tree and the historically named cultivar, and congruence is examined. This system is applied at two levels: streamlined and comprehensive. The streamlined approach for the current DNA profile dataset efficiently determined whether each individual belongs confidently in the “Reference Panel” or is relegated to “Accessory Profiles” pending further evidence. A Reference Panel was assembled of hundreds of U.S. heirlooms (and more than a thousand close relatives from other regions). The comprehensive approach involves attention from a transdisciplinary tribunal of experts who carefully weigh evidence that an apple individual – often a proposed new discovery of an otherwise lost heirloom – is indeed a historically named cultivar. As MyFruitTree accumulates more DNA profiles submitted by cultivar collection managers and apple enthusiasts nationwide, the cultivar name assignment system is being applied to unidentified trees with strong provenance evidence of cultivar status. Establishing accurate cultivar identities of valued trees via DNA profiling is providing the critical foundation for a coordinated national effort to sustain preservation and utilization of apple crop diversity.
U.S. heirloom apple cultivars are an underutilized and threatened resource, that DNA fingerprinting can help save and remobilize. These heirlooms are old cultivars that were named, clonally propagated, and distributed more than a century ago. Many heirloom cultivars have great historical value, some are still grown commercially, and others could be reintroduced to enhance rural prosperity and diversify options for consumers. While some heirloom cultivars are ancestors of modern cultivars, many others could be valuable for future breeding. However, most heirlooms have been long neglected, and thousands once documented are already extinct. Before more heirlooms disappear forever, mystery trees need to be distinguished from known cultivars, identified, and adequately preserved. Leaf samples for more than 2000 apple trees in collections, national heritage sites, old orchards, and backyards across the U.S. were crowdsourced from about 150 “MyFruitTree” submitters and DNA fingerprinted using KASP genotyping with 48 SNPs. The cultivar identity or uniqueness of each tree was determined by comparing obtained DNA profiles to a previously developed dataset of several thousand apple cultivars and individuals. Trees with replicates were prioritized into five categories according to several criteria for likelihood of representing heirloom cultivars. After removing duplicate samples, poor genotypic data, and non-apple samples, about 60% (1202) of samples were identified as cultivars and most were heirlooms. Of the unidentified samples, 85% (665) were unique, and 15% (118) of the samples represented replicated trees. We found five “Priority 1” trees (filled cultivar pedigree gaps or from three U.S. regions) and four “Priority 2” trees (detected in two regions). Hundreds more trees likely representing unknown heirlooms were also identified in single regions. Collaborators across the country, including historians and citizen scientists, can now closely examine the highest priority trees to uncover their historic cultivar names, while ensuring they are propagated so that they are preserved and valued once again. As more old apple trees are DNA fingerprinted, it is expected that current “unique” DNA profiles will be replicated in the same or other regions. Replicated trees must represent propagated, valued, and likely named cultivars, increasing the opportunities to rediscover lost heirlooms.
Peach trees require quantitative exposure to winter chilling (chilling requirement, CR) for spring bloom. The chill accumulation time points are determined using weather data of air temperatures between 32 and 45 ºF, using various calculation methods such as the simple chill hour (CH) method or more sophisticated methods like Utah and Chill portions (CP) that account for negations of chilling due to warm weather during the dormancy. All these methods rely on air temperature and do not consider the soil temperature during the dormancy and its effect on the tree’s perception and account for chill accumulation. Peach flowers and developing fruit are highly sensitive to freezing temperatures and are killed following even a limited exposure. In the past decade, mild winters and early spring frosts have significantly reduced or eliminated the annual peach production in the southeast U.S. Low-chill winters have become increasingly common in the southeastern peach-producing regions, and when followed by warm springs, result in early bud break and early flowering, increasing the risk of crop loss to frost. Due to a replant issue caused by Armillaria root rot, almost all acreage under the new peach orchards in the southeast, including South Carolina, are planted on berms adopting root collar excavation as a method to extend the life of orchards on infested soil. We observed significant differences between the air temperature and temperature of undisrupted soil and soil within berms at various depths (3, 6, 12 and 18 in) during dormancy. The effect of observed temperature differences on ‘Cresthaven’ tree chill accumulation calculation and transition between endo- and eco-dormancy stages was investigated by collecting vegetative bud and root tissue from all four depths at six chill hour time points (400, 500, 600, 700, 800 and 900). Preliminary data show significant gene expression differences between bud and root tissue and different gene expression profiles related to the chill accumulation in each tissue. Detailed analyses of the gene expression profiles between the tissues at the different chill accumulation stages and their effect on chilling and heat accumulation, bloom time, and the transition between the dormancy stages in peaches will be discussed.
Since no Huanglongbing (HLB)-resistant citrus cultivar is available in the world, selection of elite natural mutants of commercial citrus for HLB-resistance/tolerance becomes a much more appealing breeding approach, especially in HLB-epidemic regions. In this study, we have selected and evaluated more than 30 citrus mutants from commercial citrus varieties in the past eight years in Florida. After greenhouse and field trials with high HLB disease pressure, we have identified several citrus lines with improved HLB-resistance/tolerance, which can be released or used for large scale of field trials. Our analyses of these lines have revealed that citrus resistobiome plays a role in the HLB resistance/tolerance, which involves a plant virus that can enhance plant resistance and illustrated the pursuit of breeding for biocontrol and a healthy microbiome. Meanwhile, we revealed that transposons have driven the selection and diversification of sweet orange (SWO). We identified six transposon families with up to 8900-fold activity increases in modern sweet orange cultivars tracing back to a common ancestor ~500 years ago. Notably, these six families of transposons contribute significantly to the formation of major cultivar groups, with frequent independent activations or accelerations observed in the breeding history of SWO. We will discuss the molecular mechanisms underlying the improved HLB-resistance, especially how the resistobiome plays a role in the improved HLB resistance/tolerance, and how to implement this new approach by utilizing and expanding the breeding of citrus resistobiome for the control of citrus HLB.
Avocado (Persea americana) is renowned for its high nutritional value and its global consumption is steadily increasing. Currently, only a few cultivars with limited genetic variability are cultivated, and there is a need for developing new avocado cultivars with enhanced horticultural, fruit quality and nutritional traits as well as resistance to diseases and pests. Application of marker assisted selection can significantly accelerate breeding new avocado varieties, which can take 15 - 20 years using traditional breeding methods. Towards the application of molecular markers in avocado breeding, in this report, genome-wide association studies (GWAS) of nine fruit quality traits of a diversity panel of 110 avocado accessions were explored using 4,706 high-quality single nucleotide polymorphisms (SNPs) using multiple models. In addition, genetic diversity and population structure were also investigated, which unveiled three main populations corresponding to the three major avocado botanical races representing Mexican, West Indian, and Guatemalan ecotypes. Phylogenetic study and quantitative genetic analyses suggested a closer relationship between the Guatemalan and West Indian races compared to the Mexican race. Genome-wide association study revealed twelve markers distributed over eleven genomic regions strongly associated with fruit quality traits including fruit color, shape, taste, and skin texture. Annotation analyses of these genomic regions revealed candidate genes affecting these traits. These findings contribute to a comprehensive understanding of the genetic composition of avocado germplasm, which will be useful for identifying genes governing fruit quality traits as well as for accelerating breeding and parent selection efforts in the avocado breeding pipeline.
Avocado (Persea americana) is the major fruit cultivated in southern Florida counties with a value exceeding 20 million dollars annually. While production in other regions is dominated by the Hass cultivar, south Florida is unique in production of the increasingly popular, green-skinned varieties. Recently, the avocado industry in South Florida has been devastated by laurel wilt (LW), an insect-disease complex spread by Raffaelea lauricola (Rf), a fungal symbiont of redbay ambrosia beetle (Xyleborus glabratus Eichhoff). Current management practices including prophylactic fungicide injections, tree rejuvenation and ambrosia beetle population reduction are costly and onerous . Unfortunately, no mature avocado trees tolerant to LW are available to growers and genetic mechanism of LW tolerance observed in some avocado seedlings is unknown. In this study, a chromosomal genome of avocado cv. ‘Simmonds’, a ‘West Indian’ (Lowland) ecotype was assembled from Pacific Biosciences HiFi reads. The genome assembly contained 451 scaffolds spanning 98.89% of the avocado genome, a N50 of 82.34MB and a BUSCO score of 95%. This assembly served as a reference genome to generate 9198 genome wide single nucleotide polymorphisms (SNPs) using genotyping by sequencing (GBS) reads of a germplasm collection comprising 80 accessions of three avocado ecotypes (Mexican, Guatemalan and West Indian) and 18 novel hybrids exhibiting seedling tolerance to LW. Phylogentic analyses revealed three major clusters with majority of LW tolerant seedlings clustering amongst Hass derived hybrids as well as cultivars belonging to Mexican and Guatemalan ecotypes such as 'Winter Mexican', and 'Ettinger'. This work provides genomic resources for characterization of genetic tolerance of LW in avocado germplasm collections and is a significant step in developing LW tolerant hybrids to support local avocado industry.
The scientific and commercial interest in pomegranate (Punica granatum L.) cultivation has increased noticeably during the last two decades. Because of the high concentration of bioactive compounds and its nutraceutical properties, pomegranate has been defined as a super food. The consumption of pomegranate juice or arils has been related to several possible benefits on human health. Recent studies have highlighted an antioxidant and anti-inflammatory activity of this fruit which seem to prevent cardiovascular, neoplastic, neurological, metabolic, and intestinal disease. The areas of cultivation of this crop are exposed to current and future challenges like long term-drought conditions and invasive pests and diseases. Increasing the biodiversity of pomegranate has been proposed has the main strategy to reduce the risk of food system vulnerability related to monoculture and the valorization of marginal land. In order to develop advanced genetic tools to improve pomegranate breeding program efficiency we present the de novo sequencing of the ‘Wonderful’ pomegranate genome. DNA isolated from diploid leaf tissues was sequenced using long read sequencing technology (PacBio), while DNA extracted from haploid pollen grains was sequenced using short reads (Illumina). Genomic data from single haploid gamete cells were analyzed using the R package called ‘Hapi’. This allowed to infer chromosomal haplotypes obtaining a higher resolution for DNA variants detection and investigating recombination events in single gametes. Although ‘Wonderful’ represents the industry standard in the United States, several cultivars with desirable traits, such as low acidity and soft seednesses, have been identified in the national germplasm. The results of this study will provide the genomic data required to investigate differences among cultivars and create trait-gene associations. This will allow breeders to facilitate the integration of desired quality traits into new germplasm resources.
A forum for discussion of potential collaborations with regards to fruit, vegetable, and edible crops – i.e. citrus, breeding, production systems, postharvest, pomology, crop management, viticulture, etc.
Updates on Curation and Standardization of Phenotypic and Genotypic Data for Horticultural Databases - Jill Bushakra Identification Of Branched-Chain Amino Acid Derived Volatile Loci In Tomato (Solanum Lycopersicum) Using GWAS And WGCNA - Austin hart From 'Agawam' to 'Zinfandel': Fruit Quality And Metabolite Diversity In The USDA Grapevine Repository - Victoria Meakem Moving Beyond Montmorency: Exploring the Genetic Diversity of Tart Cherry - Benjamin Gutierrez Population affects growth and plant architecture in wild-collected Hydrangea quercifolia - Lisa Alexander
The Genome Database for Rosaceae (GDR, https://www.rosaceae.org/) and the Genome Database for Vaccinium (GDV, https://www.vaccinium.org/), are databases that support genomics, genetics and breeding in under-represented crops like small fruits. These fruit crops include Fragaria (strawberry), Rubus (red raspberry, black raspberry, and blackberry) in GDR, and Vaccinium (blueberry and cranberry) in GDV. Data include curated genome sequences, genetic maps, markers, QTL, genes, transcripts, germplasm, and publications, made accessible to browse, query and download through easy-to-use web interfaces and tools. One of the objectives of a 2022-funded SCRI- project ‘Advanced National Database Resources for Specialty Crop Research and Improvement’ is to collect, curate, and integrate all types of genomics, genetics, and breeding big data in easy-to-use and robust crop-specific databases. In this presentation, we summarize our progress towards curating and making available phenotype and genotype data for strawberry. We also present a strawberry Crop Ontology we have developed with input from crop researchers and breeders from North America and Europe. Public availability of phenotypic and genotypic data in GDR, GDV, and GRIN-GLOBAL will allow easy access to this data to use in genome-wide association studies. Crop Ontology will enable digital capture and trait data integration across locations and projects.
A major focus in plant breeding has been the improvement of crops through various traits that affect disease resistance and yield. However, the focus on productivity has led to an inattentiveness to other traits that specifically affect produce quality. An example of a critical fruit quality trait is its flavor, contributing to our perception of aromatic volatiles. Even at nanomolar concentrations, aromatic volatiles can be perceived by the olfactory system and influence the liking of the fruit. The focus of this study was to investigate the genetic aspect of the branched-chain amino acid (BCAA) volatiles, derived from L-valine, L-isoleucine and L-leucine, in tomato fruits. It is generally considered that these BCAA-derived volatiles contribute positively to overall liking, because these are essential amino acids required by the human diet. To identify quantitative trait loci (QTLs) affecting the biosynthetic pathway for 11 BCAA-derived volatiles, a Genome-Wide Association Study (GWAS) was conducted using a diverse and unique panel of 167 tomato accessions. The GWAS was run using the FarmCPU model in GAPIT-R, with a total of 21,893,681 SNPs, 2,735,297 INDELs, and 154 structural variants across the genome. Furthermore, a weighted gene co-expression network analysis (WGCNA) was conducted in parallel to identify modules of co-expressed genes that cluster with known genes that affect the BCAA-derived volatile pathway. A total of 113 QTLs were identified from GWAS and 3024 co-expressed genes were identified from the WGCNA. Candidate genes were screened based on annotated biochemical function, overlap within the GWAS QTLs, and gene expression in the red-ripe fruits. This led to the identification of two candidate genes, one on the long arm of chromosome 1 and another on the long arm of chromosome 11. These genes are being knocked-out using CRISPR-Cas9, and current progress is aimed to confirm the validity and function of these genes in the near-future.
The USDA National Plant Germplasm System is a network of germplasm repositories dedicated to conserving genetic diversity of crops and their wild relatives. The USDA grapevine (Vitis) repository contains 5000 unique accessions representing 36 species, and is divided between two locations: Davis, CA and Geneva, NY. While this material is available for distribution to researchers and breeders, there is currently limited characterization data to help requestors identify accessions with unique and valuable traits, particularly for fruit quality. Thus, we began a germplasm screening project to measure fruit quality traits of 481 unique accessions from both locations spanning three years (2022-2024). Fruit samples were juiced, filtered through cheesecloth, and analyzed for Brix, titratable acidity, and available nitrogen. Additionally, composition of phenolic compounds was assessed using liquid chromatography-mass spectrometry (LC-MS), and aromas were detected using solid phase microextraction (SPME) coupled with gas chromatography-mass spectrometry (GC-MS). Overall, there was a wide range of diversity in fruit quality traits across the collection. Brix values ranged from 8.5 to 30.1 (average=17.8), and titratable acidity ranged from 2.7 to 25.6 g mol-1 Tartaric acid equivalents (average=8.2). The phenolic compounds commonly detected in grape juice included the anthocyanins malvidin 3-O-glucoside and peonidin 3-O-glucoside, the hydroxycinnamic acid caftaric acid, and the flavonols quercetin 3-O-glucuronide and isoquercetin. Analysis of aromatic compounds revealed accessions that contained higher amounts of linalool, β-myrcene, and geraniol, which are associated with a “Muscat” flavor profile, while others contained methyl anthranilate, which is associated with a “Foxy” flavor profile. We hope this fruit quality dataset not only proves to be a valuable asset to researchers utilizing the USDA Vitis repository, but may also open new directions of exploration into improved grape flavor, nutrition, and quality.
Genetic diversity is invaluable to the sustainability of American horticulture. In the case of tart cherry, production in the United States is precariously reliant on a single cultivar, ‘Montmorency.’ Our research explores diverse genetic resources in tart cherry to promote utilization of high quality and locally adapted cultivars for plant breeding and improved production. Tart cherry nutritional quality is of particular interest to consumers. The United States Department of Agriculture Tart Cherry collection in Geneva, New York maintains 100 cultivars of tart cherry, including their wild relatives. Over a five-year period, we assayed fruit quality traits, including Brix, titratable acidity, and phenolic content. Total soluble solids (TSS) ranged from 10.9 to 20.7% (average=14.8%) and acidity (TA) ranged from 5.3 to 32.1 g/L (average=16.3%). The sugar/acid ratio ranged from 3.7 to 27.6 (average=10.2). Individual fruit weight ranged from 0.2 to 8.6 g (average of 5.0 g) and pit weight percentage ranged from 6-32% (average=11%). Total anthocyanin content varied from 75.2 to 3760.0 μg/g, with an average of 771.4 μg. We also evaluated bloom phenology over a three-year period. The distribution of bloom ranged from 56.7 to 134.4 GDD, with an average value of 86.0 Growing Degree Days (GDD ). ‘Montmorency’ bloom was above average with values around 95.8 GDD. Sweet cherries (63.5 – 90.6 GDD) tended to bloom much earlier than tart cherries (64.9 –118.0 GDD) and P. fruticosa, the wild progenitor of the tart cherry, bloomed the latest with a range of 85.3 to 134.4 GDD. For Brix, acidity, and phenolic content, ‘Montmorency’ falls significantly below average, though it has a balanced sugar/acid ratio. ‘Montmorency’ is lacking in anthocyanin content which is increasingly relevant for the juice industry. It tends to bloom later than other tart cherries evaluated, though there are some more extreme late bloomers. This data will be available through GRIN-Global, the USDA germplasm database to facilitate future research and breeding.
Hydrangea quercifolia, oakleaf hydrangea, a flowering shrub native to woodlands of the southeastern United States. Oakleaf hydrangea has immense ornamental potential with four-season interest, including traits like showy panicles, striking foliage textures, red fall color, and exfoliating bark. Cultivars are often derived from wild selections either directly or only a few generations removed. Full genetic and phenotypic variation has not been evaluated for the species, and little is known about the diversity in horticulturally important traits for oakleaf hydrangea. For this study, growth and plant architecture of wild-collected oakleaf hydrangea seedlings were observed at the Otis L. Floyd Nursery Research Center in McMinnville Tennessee over a 3-year period. Seedlings from 14 populations of oakleaf hydrangea spanning the species’ native range were planted in a randomized complete block design containing six blocks and nine replications per block. Two-way ANOVA was used to partition variation in height, width, growth rate, and number of stems into sources attributable to block, population, and block × population. There were significant differences among populations for growth rate, size, and number of stems in all years. Southern populations were smaller than northern populations and showed a slower growth rate. Genetic and phenotypic variation shown among populations will guide conservation efforts and supplement breeding efforts for oakleaf hydrangea.
