Source: N Y AGRICULTURAL EXPT STATION submitted to
VITISGEN2: APPLICATION OF NEXT GENERATION TECHNOLOGIES TO ACCELERATE GRAPEVINE CULTIVAR DEVELOPMENT
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
EXTENDED
Funding Source
Reporting Frequency
Annual
Accession No.
1013060
Grant No.
2017-51181-26829
Project No.
NYG-632586
Proposal No.
2017-03198
Multistate No.
(N/A)
Program Code
SCRI
Project Start Date
Sep 1, 2017
Project End Date
Aug 31, 2022
Grant Year
2017
Project Director
Reisch, B. I.
Recipient Organization
N Y AGRICULTURAL EXPT STATION
(N/A)
GENEVA,NY 14456
Performing Department
Horticulture
Non Technical Summary
Recent analyses have emphasized the economic and environmental importance of developing new grape cultivars with high fruit quality (FQ) and resistance to powdery mildew (PM). An Advisory Panel of table, juice, raisin and wine grape industry members concurs that high-quality, PM-resistant cultivars are a top priority. The proposed project (VitisGen2) will i) expand on VitisGen progress in developing novel economic, phenotyping and genetics knowledge and tools related to new grape cultivars, and ii) translate these and previous VitisGen innovations into new applications for improving grape breeding programs and managing existing vineyard plantings. The Economics team will evaluate the consequences of introducing new grape traits, including impacts upon cost, yield, revenue, profit, pesticide use, and the environment. The Trait Evaluation team will develop novel high-throughput methods, and apply these along with proven approaches to characterize a range of phenotypes, such as PM resistance durability and undesirable fruit qualities, as well as locally important traits. The Genetics team will couple phenotyping results with high-resolution genetic maps, which can be combined with genome assembly and RNA-Seq analyses to develop inexpensive, high-resolution markers spanning key genes. Finally, the Outreach team will communicate scientific opportunities and discoveries, and provide stakeholders with knowledge of the benefits of adopting new high-quality PM-resistant cultivars along with new tools for characterizing PM and FQ in their existing plantings. The proposed work utilizes plant breeding and genomics approaches to improve grape characteristics and enhance the economic and environmental sustainability of grape production.
Animal Health Component
0%
Research Effort Categories
Basic
(N/A)
Applied
50%
Developmental
50%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2011139108130%
2041139100020%
2121139110220%
2031139102010%
6011139301010%
9031139303010%
Goals / Objectives
Powdery mildew (PM) resistance consistently ranks as a top research priority facing the U.S. grape industry, based on more than a dozen stakeholder meetings focused on research priorities since 2005 and numerous industry surveys. PM is the most significant grape disease in California - and likely worldwide - in terms of expenses for control and losses in quality and yield. In an international survey of grape scientists, PM was statistically significant in being both the most important and the most genetically tractable of twelve key traits. However, breeding for PM resistant cultivars via traditional methods frequently results in negative fruit quality trait introgression from wild Vitis. Winemakers report that poor sensory attributes and color are major challenges to using interspecific hybrids, particularly the presence of high acidity, low tannin and color stability, and off-aromas. To address the need for PM resistant cultivars with desirable fruit quality, the long-term objectives of the project are listed below:1. Technological Innovations Driven by Trait Economics.Integrate genome-wide data with innovations in phenotyping powdery mildew (PM) resistance, and table and wine grape quality for genetic characterization of high-priority traits. Specific goals focus on gene discovery and trait assays: de novo annotation of relevant wild genomes; RNASeq of all VitisGen parents; low-cost AmpSeq marker implementation; automated PM quantitation without staining; multiple high throughput phenotyping screens for key fruit quality traits, and identification of candidate genes for PM resistance and fruit quality.Complete and disseminate economic analysis of several key agronomic and quality traits to drive research and breeding.2. Knowledge Extension and Application.Incorporate technological innovations and economics-oriented priorities in the generation of grapevine seedlings in breeding programs and the selection of elite breeding lines. Publicly release grapevines, pollen, and/or seed lots with various combinations of RUN1, REN1, REN2, REN3, REN4, REN6, REN7 and REN10 PM resistance.Demonstrate the impact of VitisGen advances to grape growers, enologists, and specialty crop researchers.
Project Methods
Methods for economic analyses to inform grower and breeder decisions: Market models will be parameterized with micro-level measures of gains to individual producers (e.g., cost savings, yield improvements, other advantages) or consumers (perceived value of and willingness-to-pay for specific quality traits or other product or process attributes). A significant part of the work is to estimate these micro-level values. Some measures can be inferred by analysis of observations of consumer willingness-to-pay at retail or buyer willingness-to-pay at wholesale.Choice experiments will be used to estimate the value of specific cultivar traits to consumers and producers using methods exemplified by Weaber and Lusk (2010), Norwood and Lusk (2011), and Lusk and Norwood (2009). Table grapes show the greatest promise for work on assessing consumer attitudes.Methods for centralized genotyping and sequencing: (1) continue the high-throughput generation of genetic maps for new VitisGen2 core and locally maintained families, continue to translate trait-associated markers and candidate genes into AmpSeq marker panels for cross-family validation, and expand efforts to provide SSR and AmpSeq marker data for application in breeding programs. (2) Breeders will make crosses consistent with local program goals, and incorporate marker technologies, especially in relation to disease resistance and fruit quality. New fruit quality markers will be implemented and resistance loci will be stacked. Markers employed will transition from SSR markers to AmpSeq haploblocks spanning trait loci, including markers associated with candidate genes. (3) Genomes of two wild species and a table grape cultivar will be assembled following PacBio sequencing, and all the parental genotypes of mapping families used in VitisGen1 and VitisGen2 will have RNASeq references prepared. These resources will enable reanalysis of GBS markers lost using alignment to the current 'PN40024' reference, and projection of markers onto relevant, annotated genomes. (4) RNASeq expression analysis will be completed for extreme phenotypes and relevant tissues pertaining to PM resistance, fruit chemotypes, and local phenotypes. Replicated bulked segregant RNASeq analyses will be used for cost effectiveness in some cases, such as when only one trait is of interest.Methods for powdery mildew (PM) resistance phenotyping: (1) This project will focus on the application of hyphal transect data, optimizing semi-automated image collection and automated image analysis for hyphal transect data. Optimization will require evaluation of different optics, sensors, and robotics compatible with growing PM in monoxenic culture on hundreds of full-sibling progeny subsamples per environment.(2) Assuming the budgeted personnel and current throughput with staining and microscopy rating of hyphal transects, we can screen 6 novel resistance sources, in three independent experiments each with more than 150 F1 progeny and 8-fold replication per experiment.(3) Stacks of resistance loci will be tested to identify complementary genes for improved durability, following our methods in Feechan et al. (2015). The initial work on this objective will focus on combinations of RUN1, REN1, REN2, REN4, REN6, and REN7, using full-sibling vines having from zero to five of these alleles.(5) Candidate gene discovery by RNASeq analysis will focus on REN1, REN2, REN3, REN4, REN6, REN7, and REN10, with the null hypothesis that these involve NB-LRR genes induced by 24 h post-inoculation.Methods for fruit quality phenotyping: For analyses at the FQ/Wine Center, sample preparation and analytical pipelines will follow VitisGen1-validated protocols. For each individual, replicate 100 g frozen berry subsamples will be thawed and homogenized. A portion of the juice will be retained for analysis of basic grape parameters (pH, Brix, TA, avg. berry weight) using common protocols. Homogenate will then be aliquoted into an appropriate number of vials to facilitate analysis across multiple platforms, CaCl2 added as necessary to interrupt further enzymatic activity, and frozen at -20 C until further chemical analysis. All analyses will be performed in replicate.(1) Marker Discovery in Time-Resolved Samples and Replicated Sites. Fruit will be sampled from two VitisGen1 families ('Horizon' × Illinois 547-1; and V. riparia '37' × 'Seyval' F2) at three time points (40 d post bloom, 25 d post-veraison, 50 d post-veraison). We will begin to address the genotype × environment interactions affecting development-related responses by the analysis of the V. riparia '37' × 'Seyval' F2 family in two geographic locations.(2) Development and Validation of Markers for Fruit Chemistry. Fruit will be available from some new VitisGen2 families in 2017, and from all individuals by 2018. Thus, Year 1 will be used to screen a subset of fruiting individuals for chemotyped traits to select families for full phenotyping in Years 2-4. We expect that the two core VitisGen2 families will segregate for each fruit quality trait category and be used for validation. Two VitisGen1 families ('Cabernet Sauvignon' × 'Norton' and MN1264 × MN1246) previously shown to segregate for key traits will also be included.(3) Development and Validation of Markers for Table Grape Qualities. Individual vines will be managed to ensure equal number of clusters per vine to reduce variability caused by disparate crop loads in two families (B37-28 × C56-11 and C81-227 × Y135-43-04) with approximately 280 individuals each. Phenotyping of rachis architecture and berry shape and size will utilize image-based field evaluation of individual clusters, with and without berries attached. Images will be normalized using a standard color card and metric ruler included with each sample, and analyzed using macros implemented within ImageJ. Post- veraison clusters will also be weighed. When sufficient berry numbers are available, clusters will be measured for berry texture (Zwick ZN250 Analyzer) and juice production. Phenotypic data will be used by the Genetics Team to identify QTL. Genetic analysis of red color stability in table grapes will be evaluated using one family phenotyped for hue and chroma as well as a subjective rating of red color of mature fruit for three seasons.Evaluation Plan: The Project Manager and an appointee from the Advisory Panel will develop and administer annual project evaluations. The evaluations will solicit written feedback from all project participants, including the Advisory Panel, and seek input on logistics, communications, and progress from the perspectives of both scientific and industry stakeholder needs. The evaluations will also provide an opportunity for suggestions on how to better deliver information for implementation. The Project Manager and Advisory Panel appointee will benchmark and track progress using the Logic Model (see project narrative, Executive Summary), the Evaluation Plan (see project narrative, Table 7), and review of annual team reports and summary tables. Based on the evaluations, the Project Manager and the Advisory Panel appointee will generate and distribute to participants an annual evaluation report, and alert the Project Director to any critical deficiencies or unanticipated opportunities identified. With the assistance of the Executive Committee and guidance from the Advisory Panel, the Project Director will develop an action plan to address the deficiencies and pursue opportunities to meet and/or exceed project goals. Further details can be found in Table 7, page 24 of the project narrative.

Progress 09/01/20 to 08/31/21

Outputs
Target Audience:The range of audience reached by the project this year include students at all levels, researchers/scientists, breeders, vineyard and winery managers, and policymakers. The project has directly and indirectly impacted wine, table, and raisin grape growers and producers around the participating regions - Minnesota, North Dakota, South Dakota, Montana, California, New York, other US states and the wider international grape growers and consumers. Changes/Problems:The supply chain issues, and shipment delays propagated by the COVID-19 pandemic impacted the speed and efficiency of samples management, processing, and DNA sequencing. Extracted DNA samples were held/trapped at distribution centers in Europe and the US for days to weeks due to the persistent shipping issues to and from Intertek, Sweden. The returned DNA was of low quality, but the efforts of the Cornell University Sequencing staff were able to save the samples and return usable data for rhAmpSeq markers. As a result, the return of MAS data was delayed, though analysis and selection outcomes have been delivered to breeding team members. Supply chain issues also slowed RNAseq samples isolation and metabolomics attempts by 6 months or more, but processing is starting to catch up with sample backlogs. Hiring freeze across many institutions due to COVID-19 pandemic affected the employment and effective management of personnel to work on the project. The pandemic regulations delayed planned visits by our engineering colleagues from Spain who had planned to improve robotic performance, slowing some aspects of the automated imaging work by two years. The COVID-19 related policies including maintaining physical distancing caused changes for most laboratory experimentation including disease resistance phenotyping, fruit quality, and other experiments that require multiple persons in a given space. This led to shifting our priorities from laboratory activities to field data collection and data analyses. What opportunities for training and professional development has the project provided?Our efforts to provide training and professional development continue to yield results. At USDA, Geneva, and Cornell University, we trained several postdocs, graduate students, and technicians on the use of imaging robots and neural network analysis. An undergraduate student received training via the USDA's 1890 National Scholar program for remote training on grapevine pathology and computer vision analysis and will return next summer for in-person research. An undergraduate researcher examined the genetics of flower timing, cluster imaging, and other aspects of grapevine breeding under our internship training program. Five undergraduates and one graduate student received training in chemical trait evaluation on grape accessions at the Fruit Quality and Wine Center throughout the academic year (Sep 2020 - May 2021). A postdoc received training in metabolomics and RNASeq data analysis. Another graduate student received training in the operation and maintenance of a GC-MS instrument for the quantification of grape-derived volatiles. Four graduate students and one postdoc received training trained in leaf material collection for DNA extraction, field phenotyping for disease (powdery mildew, downy mildew), phenology (bud break, bloom, veraison, wood acclimation), and fruit composition at NDSU. The Trait Economics team trained two research assistants and a student on trait economics data assembly and analysis. A postdoc developed capacity on integrated GBS-rhAmpSeq genetic map development, phenotypic data curation, and manuscript writing while an undergraduate received training in viticulture and phenotyping vine growth traits at SDSU. An assistant project scientist and one research specialist developed capacity and built new tools in bioinformatics for the advancement of grape genomics at UC Davis. A deaf-blind PhD student has contributed effort to the project since its inception and will defend her PhD in April 2022. One postdoctoral scientist on the project already found placement with a private breeding company, and another postdoc obtained a faculty research position. One technician was recruited to a PhD program at Arizona State University. One undergraduate viticulture student accepted a job in education, and another undergraduate viticulture student began a PhD program in Horticulture. How have the results been disseminated to communities of interest?Communications about project accomplishments, developments, and personnel were continuously carried out using the project website, web broadcasts, seminar and conference presentations, scientific and non-scientific meetings, publications (trade publications, scientific publications), and social media platforms. Due to the restrictions related to the COVID-19 pandemic, many of our scientists, students, and technicians participated in a mix of meeting formats, in-person and virtual. A few of the notable meetings and conferences including the Agricultural and Applied Economics Association (AAEA) AAEA & Western Agricultural Economics Association (WAEA) Joint Annual Meeting, Austin Texas; the Department of Agricultural, Food, and Resource Economics seminar series, Michigan State University; Ohio Grape and Wine Conference; the North Dakota Grape and Wine Association meeting; the Montana Grape and Winery Association Meeting; North Dakota State University Grape Germplasm Enhancement Project; the American Society for Enology and Viticulture National Conference; the American Society for Enology and Viticulture- Eastern Section Annual Conference; the North Dakota Academy of Science 112th Annual Meeting; the 36th Annual Plant Science Graduate Students' Symposium, University of Saskatchewan; the North Dakota State University- Gamma Sigma Delta Symposium, the North Dakota State University - Graduate Student Council Research Symposium and the Local Foods Education Center, Brookings, SD. The robotic phenotyping aspects of the project were featured in the podcast series of the American Phytopathological Society: Plantopia (see: https://www.plantopiapodcast.org/19). The USDA, Geneva/Cornell University hosted visits by the Cornell University CALS Dean and several NY state congressional representatives to discuss the importance of the multi-institutional collaborative research. Many scientists in the project participated in field visits, field days, teaching, and student training. The extension and outreach arm of the project hosted several webinars, trade publications, Research in Plain English write-ups, blog posts, etc. using the VitisGen2 website, other web services, and social media to disseminate information about the project. What do you plan to do during the next reporting period to accomplish the goals?The genetics team plans to focus on finishing up genetic mapping efforts across the project and publishing project outcomes on MAS and genetic mapping of QTL. The team will sustain the efforts on RNAseq analysis from different resistant sources to powdery mildew including Vitis amurensis 588631, Tamiami B37-28 (REN11), V. romanetii (REN4), and RENstack1 (RUN1/REN1/REN6/REN7) and a novel qualitative downy mildew resistance from V. x doaniana 588149 as well as RNAseq analysis for fruit quality projects. There are plans to complete the sequencing and the assembly of the various sources of PM resistance: V. doaniana 588149, Y540-6-15, Tamiami (REN11+), MN1264 (REN3+/ REN9+/ REN10+), and Y522-123-15 (RUN1+/REN1+/REN4+), coupled with shotgun sequencing of phenotyped recombinants, to define the newly discovered loci associated with PM resistance. Additional germplasm focused on breeder-driven priorities for resistance to phylloxera and downy mildew will also be subject to this strategy to characterize resistant loci. Regarding the characterization of the genetic basis of V. piasezkii's PM resistance, DNA-seq data of eight F1 accessions from a breeding population, 11373, will serve to distinguish and reconstruct the two haplotypes of REN6 and REN7 loci in the chromosome-scale diploid assembly of V. piasezkii DVIT2027 (REN6+/REN7+). The genomic regions composing the two loci will be identified among the genome assemblies of six F1 accessions and then compared with V. piasezkii's haplotypes to determine REN6- and REN7-associated genes, short polymorphisms, such as single-nucleotide polymorphisms (SNPs) and short insertion-deletion (INDELs). From these analyses, we expect to identify candidate PM resistance-linked genes and to develop markers for marker-assisted selection (MAS). To better understand the similarities and differences across diverse sources of the REN1 locus, we will compare the REN1 haplotypes from the genomes of the six V. vinifera spp. vinifera: Husseine (REN1+), Karadzhandal (REN1+), Khalchili (REN1+), Late Vavilov (REN1+), Sochal (REN1+) and Y315-43-04 (REN1+), and four publicly available V. vinifera spp. sylvestris genomes: DVIT3351.27 (REN1+), DVIT3603.07 (REN1-), DVIT3603.16 (REN1+), and O34-16 (REN1+). An additional goal for the genetics team is to continue to develop methods to incorporate the rhAmpSeq data into the practical haplotype graph structure used by other breeding programs in the development of genomic selection tools. The team has partnered with the Breeding Insight project to genotype the existing USDA germplasm to identify specific genotypes which carry key haplotypes in the repository system. In general, most of the breeding centers will continue to maintain their existing mapping populations and focus on finalizing data analyses from different projects and to publish relevant research results for both scientific and non-scientific audiences. Data analysis on mapping families using recently obtained data from the previous year(s), the implementation of marker-assisted breeding with additional loci, in particular, malic acid and foliar phylloxera resistance, and the development of new genetic maps for populations for fruit quality and Ren10 powdery mildew resistance locus will continue at the University of Minnesota. Following the phenotypic and genotypic selection of individuals as future parents (with powdery mildew resistance incorporated), additional crosses will be conducted in 2022 to bridge the allele(s) into more environmentally durable backgrounds for the North Dakota climate, fruit composition and phenology data analyses will be conducted for previously submitted interspecific hybrid families, marker-trait association refinement will be conducted, and Greenhouse-grown progenies of interest will be phenotyped for traits of interest in the winter of 2021-2022 at NDSU. Validation of markers for the selection of anthocyanin monoglucoside red wine types, field test selections from MAS carrying combinations of new Amurensis-derived DM resistance loci, and the development of models for efficient foliar traits assessment using a hyperspectrometer, QTL mapping, and genetic studies of foliar-related ions and compounds will continue at Cornell University. In collaboration with the powdery mildew team, PM, and DM data will be analyzed at SDSU. Also, fruit quality analysis of F2-VRS (V. riparia x Seyval) from plots in CA and SD will be completed and a study of the genetic architecture of the root system will be published. The Trait Economics team will finalize the analysis of the consumer survey data, publication, and dissemination of results to the public as well as relevant academic audiences while the PM team will focus on the publication of results and making our discoveries and technologies widely available. The Fruit Quality team will complete phenotyping analyses of 2019 and 2020 samples, integrate, and publish metabolomic and transcriptomic data from a time-resolved sampling of Vinifera, Riparia, and Cinerea genotypes with different malate behavior during ripening. Also, the team will perform metabolite and transcript analyses on fruit of different grape species stored under normoxic and hypoxic conditions to test the hypothesis that differential response to hypoxia can explain variation in malate behavior among grape species as well as complete RNAseq analysis of diverse genotypes to confirm the relevance of candidate transcripts in explaining differences in malate metabolism. Finally, the Extension team will publish about the release of RenStack vines to the public domain, preferred table grape attributes, and the influence of breeding methods (conventional, gene editing) on consumer acceptance among other topics.

Impacts
What was accomplished under these goals? With the addition of ~9000 new samples, the total number of grape seedlings processed with the rhAmpSeq sequencing technology developed by the VitisGen2 project, comes up to ~71,700 samples. The project has provided DNA services to 19 breeding and genetics programs in the public and private sectors, both domestic, and international, covering bunch grapes and muscadine, including USDA grapevine germplasm. Fifty-eight quantitative trait loci (QTL) associated with important grape attributes have been identified. This has promoted the implementation of marker-assisted-selection (MAS) across breeding programs for disease resistance (RUN1, REN1-11, RPV), muscat flavor, seedlessness, diglucosides, flower sex, and seed trace size selection. The implication is that breeders can now easily discard seedlings with undesirable traits based on marker data at the seedling stage, thereby, reducing cost and saving resources while improving selection accuracy. In 2020, the genetics team opened a fee-for-service opportunity for public and private breeders not funded by the project. A total of >2000 markers has been assessed for ~6,000 seedlings in 2021. Also, the project has continued to generate genome assemblies and whole-genome sequencing of powdery mildew (PM) resistance sources using emerging technologies. In prior years, the capacity to process and image thousands of leaf disk samples per day was developed, but the custom trays designed to hold the disks were too expensive. This year, an injection mold that enables inexpensive mass production of the custom trays was designed and procured in partnership with Printersys. Also, a saliency mapping algorithm that improves the accuracy of PM quantification and shows biologist which pixels the algorithm sees as PM was refined for image analysis. This increases confidence in the results and improves QTL detection. Technological Innovations Driven by Trait Economics: This year, the rhAmpSeq markers have been sequenced for ~5,600 samples with an additional 3,600 samples in the sequencing queue. In the past year, collection of plant material and isolation of RNA was improved, and tissue samples have been collected for evaluating differences in fruit quality and disease resistance. Research efforts including linkage map construction, QTL analyses and seedling selection were supported in the different breeding programs using previously validated markers. Genome assemblies of Single Molecule Real Time (SMRT) DNA sequences (Pacific Biosciences (PacBio)) of genetic sources of PM resistance was completed for Late Vavilov (REN1+), V. vinifera ssp. vinifera Y315-43-04 (REN1+), the two parents of Illinois 547-1 (REN2+): V. rupestris B38 and V. cinerea B9 (REN2+), and Horizon (REN3+/REN9+). Also, the gene annotation of V. piasezkii DVIT2027 (REN6+/REN7+) has been completed and the chromosome-scale diploid genome assembly is ongoing. Whole-genome sequencing of V. doaniana 588149 was sequenced using highly accurate long-read sequencing (HiFi; PacBio). SMRT DNA sequencing (PacBio) of Tamiami (REN11+; parent of B37-28) is ongoing. HiFi library preparation of Y540-6-15 (V. amurensis 588631 x Valley Pearl) is ongoing as well as two additional accessions: MN1264 (REN3+/ REN9+/REN10+; parent RenStack2) and Y522-123-15 (RUN1+/REN1+/REN4+; parent RenStack2 (GE1875)). Illumina DNA sequencing (DNA-seq) of two V. vinifera backcross lines, e6-23 (RUN1.2b+) and 08391-29 (RUN2.2+), was performed to help distinguish and reconstruct the two haplotypes of Run1.2 and Run2.2 loci in the chromosome-scale assembly of M. rotundifolia Trayshed. Vineyard images were collected from 6 mapping families to support vineyard image-based analysis of diseases and phylloxera. A low-cost QR code system attached to trellis wires was implemented for separation and automated identification of vines in images. Through an outside collaboration, we coordinated satellite imaging, proximal imaging, and traditional expert ratings. Two mapping families were converted to vertical shoot positioning for improved imaging of fruit clusters. Powdery mildew, downy mildew (DM), and phylloxera resistance assessments in the vineyard were conducted on ~480 genotypes for the validation of image-based vineyard evaluation. Three large-scale, laboratory phenotyping experiments were executed for grapevine PM or DM for genetic mapping, marker validation, resistance genes stacking, fine mapping and race specificity testing. About 905 vines were screened to identify recombinants at 12 resistance loci, for detailed characterization of resistance biology, improved molecular markers, and identification of candidate resistance genes. Also, AmpSeq markers were applied to 1300 new Erysiphe necator samples for translation of this VitisGen1 marker technology to fungicide resistance monitoring and population genetics. Differences in responses of wild and domesticated Vitis spp to low-oxygen conditions were characterized for the 2nd year. Grapes fruits were also stored under anoxic conditions to evaluate if wild and domesticated grapes showed differential behavior in malate metabolism in response to anoxic conditions. Fruit chemistry and volatile analyses were centrally or locally analyzed across programs. Vitis spp were characterized for differences in transcript and metabolite production related to malate regulation. Using previously collected samples of wild and interspecific Vitis from the USDA-ARS germplasm collection, RNAseq analysis was performed to identify differentially expressed genes during ripening of domesticated genotypes (vinifera) with high malate degradation, and wild genotypes (cinerea, riparia) with low malate degradation. Specific transcripts that are associated with malate behavior (degradation/lack of degradation) were identified and validated with additional samples with diverse genetic backgrounds and malate behavior. Traditional local phenotyping efforts were carried out in many mapping populations for malic acid, tartaric acid, pH, and winter cold hardiness (Missouri); grape berry color, genetic resistance to Japanese beetle feeding, floral and aromatic compounds, flowers, and fruit (Minnesota); timing of flowering and disease resistance (Cornell University); Fruit composition and phenological traits as well as differential thermal analysis for winter freeze resistance in North Dakota. Hyperspectral sensing was used in a 2nd year of chemical characterization of grape leaves for further genetic, vineyard health and association studies at Cornell University. Knowledge Extension and Application: New populations were generated to incorporate the new source of PM resistance derived from V. amurensis into breeding lines of interest at Cornell University and similar objective at North Dakota State University (NDSU). Also, the seedlings from 2020 were planted into a permanent location and assessed for resistance to disease in relation to marker data at Cornell University. Embryo culture seedlings from Run1Ren1Ren4 x fruit quality crosses were reared in the growth chamber/greenhouse until ready for field transplanting in California-Parlier. These populations for future fruit quality assessment were trained in 2021. The analysis of the economics of varietal innovation using historical data on the development and adoption of table grape varieties, as well as the analysis of results from an online consumer survey, which we implemented in 2020, to evaluate consumer attitudes towards specific table grape traits, and gene-editing in fresh fruit are in advanced stages. Detailed review and analysis of foreign policies e.g., Europe, that may affect the adoption of local varietal innovations was initiated. Information dissemination of products and innovations has been sustained using various channels - trade publications, scientific journals, conferences, etc. to different public and private stakeholders.

Publications

  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Multiple independent recombinations led to hermaphroditism in grapevine Cheng Zou, M�lanie Massonnet, Andrea Minio, Sagar Patel, Victor Llaca, Avinash Karn, Fred Gouker, Lance Cadle-Davidson, Bruce Reisch, Anne Fennell, Dario Cantu, Qi Sun, Jason P. Londo Proceedings of the National Academy of Sciences Apr 2021, 118 (15) e2023548118; DOI: 10.1073/pnas.2023548118
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Zou, C., Karn, A., Reisch, B. et al. Haplotyping the Vitis collinear core genome with rhAmpSeq improves marker transferability in a diverse genus. Nat Commun 11, 413 (2020). https://doi.org/10.1038/s41467-019-14280-1
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Yin L, Karn A, Cadle-Davidson L, Zou C, Underhill A, Atkins P, Treiber E, Voytas D and Clark M (2021) Fine Mapping of Leaf Trichome Density Revealed a 747-kb Region on Chromosome 1 in Cold-Hardy Hybrid Wine Grape Populations. Front. Plant Sci. 12:587640. doi: 10.3389/fpls.2021.587640
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Teh, S.L., Rostandy, B., Awale, M. et al. Genetic analysis of stilbenoid profiles in grapevine stems reveals a major mQTL hotspot on chromosome 18 associated with disease-resistance motifs. Hortic Res 6, 121 (2019). https://doi.org/10.1038/s41438-019-0203-x
  • Type: Books Status: Published Year Published: 2019 Citation: Cantu, D. and Walker, A. M. The Grape Genome. Springer, Cham. 1 (XXVII, 367). DOI: https://doi.org/10.1007/978-3-030-18601-2. 2019. ISBN : 978-3-030-18600-5
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Alston, J.M., Gaeta, D. Reflections on the Political Economy of European Wine Appellations. Ital Econ J 7, 219258 (2021). https://doi.org/10.1007/s40797-021-00145-4
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Negus KL, Chen L, Fresnedo-Ram�rez J, Scott HA, Sacks GL, et al. 2021. Identification of QTLs for berry acid and tannin in a Vitis aestivalis-derived 'Norton'-based population. Fruit Research 1: 8 doi: 10.48130/FruRes-2021-0008
  • Type: Theses/Dissertations Status: Published Year Published: 2021 Citation: Karlene Negus, Identification of QTLs for Berry Acids and Sugar in a Vitis aestivalis-derived Norton-based Population
  • Type: Theses/Dissertations Status: Published Year Published: 2021 Citation: Kavya Tummala, Marker-assisted Selection to Determine the Introgression of Rpv3 Mediated Downy Mildew Resistance in Chambourcin X Cabernet Sauvignon Grapevine Population
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Yin, Lu, Eric C. Burkness, William D. Hutchison, and Matthew D. Clark. Lu Yin, Eric C. Burkness, William D. Hutchison, and Matthew D. Clark "Effects of Foliar Phylloxera (Hemiptera: Phylloxeridae) Infestations on Wine Grape Photosynthesis, Yield, and Fruit Quality," Journal of Entomological Science 56(4), 504-518. https://doi.org/10.18474/JES20-77
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Olson, Jack, and Matthew Clark. " Characterization of Anatomical and Physiological Effects of Variegation Mutation on Grapevine", HortScience horts 56, 10 (2021): 1251-1257, https://doi.org/10.21273/HORTSCI15929-21
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Moreira, Laise S., and Matthew D. Clark. " Embryo Rescue of Cold-hardy Table Grapes", HortScience horts 56, 9 (2021): 1059-1065, https://doi.org/10.21273/HORTSCI15850-21
  • Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Clark, MD. 2021. Panelist: Future Grape Cultivars for Eastern North America. Annual Conference of the American Society for Enology and Viticulture-Eastern Section (ASEV-ES)Virtual Conference 2021.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2021 Citation: Moreira, L., Z. Vickers, E. Treiber, Anna Andresen and M. Clark, and A. Hegeman. (2021) A procedure to investigated flavor in large grape samples. American Society for Horticultural Science (ASHS) Conference. Denver, CO. August 7, 2021
  • Type: Other Status: Accepted Year Published: 2021 Citation: Martinson, T. , B. Reisch and R. Wiepz. (2021) The central role of mapping populations in marker-assisted grape breeding: A tale of three related mapping populations at Cornell AgriTech. Wine Business Monthly, February 2021.
  • Type: Other Status: Accepted Year Published: 2021 Citation: Martinson, T, Cheng Zou, Qi Sun, Jason Londo, M�lanie Massonnet and Dario Cantu. (2021) How Genetics Determine Flower Sex in Grapevines. Wine Business Monthly, October 2021.
  • Type: Other Status: Accepted Year Published: 2021 Citation: Podolec, M. and Tim Martinson. (2021) Breeding Insight brings bioinformatics to specialty crop breeding. VitisGen2 brings technology to the table. Good Fruit Grower. November 2021
  • Type: Websites Status: Accepted Year Published: 2021 Citation: Rodger, E. (2021) Grape genetics research reveals what makes the perfect flower. Cornell Chronicle, May 2021.
  • Type: Websites Status: Accepted Year Published: 2021 Citation: Martinson, T., Podolec, M, and V. Greenlee (2021) Partnership applies genome search engine to 7000 grapevines. Cornell Chronicle, Sept 29, 2021
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2021 Citation: Noam Reshef, Avinash Karn, David C. Manns, Anna Katharine Mansfield, Lance Cadle-Davidson, Bruce Reisch, Gavin L. Sacks (2021). Stable QTL for malate levels in ripe fruit and their transferability across Vitis species. Horticulture Research.
  • Type: Journal Articles Status: Under Review Year Published: 2021 Citation: Avinash Karn, Luis Diaz-Garcia, Noam Reshef, Cheng Zou, David C. Manns, Lance Cadle-Davidson, Anna Katharine Mansfield, Bruce I. Reisch, and Gavin L. Sacks (2021). The genetic basis of anthocyanin acylation in North American grapes (Vitis spp.). Genes, Special Issue: Genetic Approaches to Improve the Flavors and Appearance of Fruits, Vegetables and Herbs.
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Karn Avinash, Zou Cheng, Brooks Siraprapa, Fresnedo-Ram�rez Jonathan, Gabler Franka, Sun Qi, Ramming David, Naegele Rachel, Ledbetter Craig, Cadle-Davidson Lance. 2021. Discovery of the REN11 Locus From Vitis aestivalis for Stable Resistance to Grapevine Powdery Mildew in a Family Segregating for Several Unstable and Tissue-Specific Quantitative Resistance Loci. Frontiers in Plant Science 12:1868 https://www.frontiersin.org/article/10.3389/fpls.2021.733899.
  • Type: Journal Articles Status: Submitted Year Published: 2021 Citation: Islam El-Sharkawy, Minkyu Park, Daniel Vera, Devaiah Kambiranda, Pranavkumar Gajjar, Lance Cadle-Davidson, and Violeta Tsolova. 2021. Chromosome-level genome sequence assembly and genome-wide association study of Muscadinia rotundifolia reveal the genetics of 12 berry-related traits. Horticulture Research.
  • Type: Journal Articles Status: Under Review Year Published: 2021 Citation: Matthew Clark, Lu Yin, Avinash Karn, Lance Cadle-Davidson, Cheng Zou, Jason Londo, and Qi Sun. 2021. Candidate resistance genes to foliar phylloxera identified at Rdv3 of hybrid grape. In review at Horticulture Research.
  • Type: Book Chapters Status: Awaiting Publication Year Published: 2021 Citation: Vezzulli S., Gramaje D., Tello J., Gambino G., Schwandner A., Barba P., Bettinelli P., Pirrello C., Angelini E., Anfora G., Mazzoni V., Pozzebon A., Palomares-Rius J.E., Mart�nez-Diz M.P, Toffolatti S.L., De Lorenzis G., De Paoli E., Perrone I., DInc� E., Zenoni S., Wilmink J., Lacombe T., Crespan M., Walker M.A., Bavaresco L., de la Fuente M., Fennell A., Tornielli G.B., Forneck A., Ib��ez J., Hausmann L., and Reisch B.I. 2021. Genomic Designing for Biotic Stress Resistant Grapevines. in Genomic Designing for Biotic Stress Resistant/Tolerant Crops. Chittarajan Kole (ed.). Springer Nature Publishing.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Karlene L. Negus, Li-Ling Chen, Jonathan Fresnedo-Ram�rez, Heather A. Scott, Gavin L. Sacks, Lance Cadle-Davidson and Chin-Feng Hwang; Seminar Title Identification of QTLs for Berry Acid and Tannin in a Vitis aestivalis-derived 'Norton'-based population, 45th Annual Virtual Conference of American Society for Enology and Viticulture-Eastern Section, July 07, 2021.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2021 Citation: Kavya Sir Tummala, Li-Ling and Chin-Feng Hwang; Seminar Title Marker-assisted Selection to Determine the Introgression of Rpv3 Mediated Downy Mildew Resistance in Chambourcin X Cabernet Sauvignon Grapevine Population, 45th Annual Virtual Conference of American Society for Enology and Viticulture-Eastern Section, July 08, 2021.
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2021 Citation: Venkateswara Kadium, Andrej Svyantek, John Stenger, Collin Auwarter, and Harlene Hatterman-Valenti (2021). Molecular Investigation of Fruit Quality in the Cold Climate Adapted Wine Grapes (Vitis spp.). American Society of Enology and Viticulture-Eastern Section
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2021 Citation: Venkateswara Rao Kadium, Andrej Svyantek, John Stenger, Sarah Bogenrief, Collin Auwarter, and Harlene Hatterman-Valenti (2020). Analysis of Dormancy Acclimation Response In Incomplete Diallel Population Representing NDSU-GGEP. North Dakota Academy of Science
  • Type: Journal Articles Status: Other Year Published: 2021 Citation: Alahakoon D, Fennell A, Helget Z, Bates T, Karn A, Manns D, Mansfield AK, Reisch B, Sacks G, Sun Q, Zou C, Cadle-Davidson L, Londo JP. Novel berry (E)-2-hexenal QTL identified in grapevine interspecific F2 population using a dense integrated GBS and rhAmpSeq genetic map.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Zou C, Gouker F, Karn A, Fennell A, Cantu D, Xu X, Clark M, Reisch B, Sun Q, Cadle-Davidson L, Londo J. Molecular Characterization of the Sex Loci in Wild and Domesticated Grapes. Plant and Animal Genome Meeting. San Diego, CA. January 12, 2020.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Anne Fennell. Cold Genetics. North Dakota Grape and Wine Association Conference, February 29, 2020, Mandan, ND.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Cantu, D. The wild side of grapevine genomics Plant and Animal Genome XXVIII Conference. January 12, 2020, San Diego, CA.
  • Type: Other Status: Other Year Published: 2019 Citation: Cantu, D. Challenges Impacting the California Wine Industry. UC Davis Chancellors Board of Advisors. April 19, 2019. Napa, CA.
  • Type: Other Status: Other Year Published: 2019 Citation: Cantu, D. Leveraging plant and microbial genomics to study grapevine diseases. Penn State University. February 25, 2019, State College, PA.
  • Type: Other Status: Other Year Published: 2019 Citation: Cantu, D. The role of domestication, breeding, and clonal propagation on genomic diversity in grapevines. Max Planck Institute for Developmental Biology. February 12 2019, T�bingen, Germany
  • Type: Websites Status: Published Year Published: 2021 Citation: 2021, July. Pacific Northwest Ag Network Wine Minute: How to Address Powdery Mildew. WSU Extension specialist Michelle Moyer addresses the use of disease-resistant varieties as a powdery mildew management option.
  • Type: Other Status: Published Year Published: 2021 Citation: Brief summary of the Journal article: Fine mapping of leaf trichome density revealed a 747-kb region on chromosome 1 in cold-hardy hybrid wine grape populations. Summary by Matthew Clark and Lu Yin.
  • Type: Websites Status: Published Year Published: 2021 Citation: January 5, 2021. Blog post. Podcast Featuring Bruce Reisch Made Top 5 Downloads on Sustainable Winegrowing with Vineyard Team in 2020, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: February 2, 2021. Blog post. Central Role of Mapping Populations in Marker-Assisted Grape Breeding, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: February 8, 2021. Blog post. Research in Plain English article on how fine mapping of leaf trichome density revealed trait location in a cold-hardy hybrid wine grape populations now available, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: June 3, 2021. Blog post. How researchers are seeking DNA sequence variations that affect micronutrient update in Vitis vinifera, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: August 6, 2021. Blog post. VitisGen2 researchers describe improved deep learning method for quantification of Grape PM at ASABE Meeting, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: August 11, 2021. Blog post. Breeding Insight brings bioinformatics to specialty crop breeding. VitisGen2 brings technology to the table, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: September 13, 2021. Blog post. New publication by VitisGen2 team describes Ren-11 powdery mildew resistance location, M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: September 13, 2021. Blog post. Researchers discover new QTLs for grape berry acid and tannins in V. aestivalis Norton and V. vinifera Cabernet Sauvignon hybrid population. M. Podolec.
  • Type: Websites Status: Published Year Published: 2021 Citation: October 12, 2021. Blog Post. Exploring the genetics behind flower sex in grapevines. M. Podolec.
  • Type: Other Status: Other Year Published: 2021 Citation: Gavin Sacks. Polish Academy of Sciences  Institute for Animal Reproduction and Food Research Seminar Series; Virtual presentation. Wild for Wine: A Flavor Chemists Perspective on Developing More Sustainable Grape Varieties for Winemakers. April 23, 2021.


Progress 09/01/19 to 08/31/20

Outputs
Target Audience:In 2019/20, the PM phenotyping laboratory (aka, the Robot Lab) remained a frequent site of outreach events focused on communicating the importance of multidisciplinary (biology, informatics, engineering) and multi-institutional research, with about 20 tours provided to scientists and the public, such as high school students interested in engineering, graduate students in plant breeding, stakeholders from grape and biocontrol industries, USDA national program leaders, congressional representatives, and economic development experts. Through the project's website (vitisgen2.org), webinars, scientific journals, trade publication articles (Wine Business Monthly, American Vineyard, Good Fruit Grower), a Twitter feed, and non-technical summaries of VitisGen2 journal articles, we reached out to a broad national and international audience of other scientists, growers, and industry leaders, as well as the public. There were notable outreaches to key industry, farmer, and consumer stakeholder groups within the host community and states of participating VitisGen2 institutions. For example, in Minnesota and North Dakota: the Minnesota Grape Growers Association; Minnesota Farm Winery Association; and the Southern Minnesota Wine Grower's Alliance. Local grape growers and winemakers in North Dakota, Montana, and western Minnesota audiences were reached this year. The scientific and the academic community equally benefit from the numerous publications in top peer-reviewed journals, seminar presentations, workshops websites, and social media posts. We offered training and education to high school students, undergraduates, graduates, post-doctoral associates, and visiting scholars locally and internationally through various programs. Our online survey tools have been important in reaching out directly to grape end-users to generate important information such as trait preferences which are necessary for making consumer-informed improvement decisions. Changes/Problems:While we did not experience any problem that warranted a change in the proposed objectives, it is important to mention the impact of the COVID-19 pandemic on the project this reporting year. Due to major shutdowns and closures of facilities across the globe in response to the pandemic, we experienced a reduction in executing planned experiments, data collection, and analyses. The closing of DNA extraction labs, greenhouses, and sequencing centers resulted in a long delay in sample collection and submission. The delays at the sequencing centers in the regular rhAmpSeq marker processing and to provide low-cost alternatives to a single-marker analysis led to the testing of KASP and SkimSeq genotyping platforms to support quality data and results to breeders for several powdery mildew resistance loci, fruit chemistry (muscat and di-glucosides), seedlessness and seed trace size, and male-sterile and female-sterile alleles. These KASP markers were evaluated at Intertek-Sweden where DNA plates were extracted. DNA extraction services and additional commercially available KASP and SSR markers were evaluated at Agbiotech in California. Due to the COVID pandemic, it was not possible this year to set up the precise experiments needed to conduct such sensitive experiments as RNAseq studies. Rather, we decided to use existing DNA to pursue a strategy for candidate gene discovery based on the alignment of shotgun Illumina sequence data to our high-quality PacBio assemblies, for seedlings with recombinations under QTL and with existing phenotype data. This will enable us to narrow in on the specific location of candidate genes, considering structural variants in the source of each trait. At Cornell, a departure of a core transcriptomic and metabolomic analyses facility staff member led to slower processing time. However, we identified an external collaborator to perform these analyses. Some breeding groups were unable to extract DNA for analysis, but others found ways to do so, though the number of genotypes analyzed was lower and the results of DNA analyses came much later. Leaf sample collections for powdery mildew analysis were delayed, reduced, or eliminated. A proposed cluster architecture experiment was delayed at USDA-Parlier. From an ambitious plan of 28 large-scale PM laboratory experiments for the 2020 growing season, we were only able to execute 5. Breeding programs could not access the vineyards to sample leaf tissue for centralized phenotyping from approximately March through June 2020, and laboratory operations remain in reduced capacity with social distancing requirements greatly restricting our abilities to conduct large experiments. In previous years we had up to seven people in one lab to set up each experiment, and now many labs are restricted to one or two people. The robotic arm would help alleviate this, but for this robot, we only have enough trays to hold 3000 samples at a time and could not have more manufactured due to the long-term COVID-19 closure of our manufacturing partner in Spain. We are developing a partnership with an injection molding manufacturer to produce hundreds of trays to alleviate this bottleneck and provide a low-cost solution for future Blackbird customers. The shutdown and de-densification requirements after re-opening in many institutions also affected the number of personnel including interns, undergraduate assistants, and contract field labor involved in sample processing and evaluation. Five international scientists who were scheduled to come for research stays had their trips canceled or delayed. Two summer undergrad students were not able to research with us in summer 2020, two new technicians had their starting dates delayed until labs reopen, and two technician positions were abolished. Employment was affected by the pandemic. Travel and contact restrictions associated with the COVID-19 pandemic prevented in-person presentations and field days that were anticipated and scheduled; rather we reverted in most cases to online presentations and electronic publications. Also, we encountered a challenge in mapping 4 newly sequenced reference genomes associated with PM resistant loci (REN1, REN2 and REN3) onto the PN40024 genome (V1 version). The BSR-seq analysis and SMRT bell libraries had non-uniform read coverage over gene bodies, especially for long CDS (> 1kbp) and a lack of coverage of the 3'. Repeating the sequencing will likely solve this issue. What opportunities for training and professional development has the project provided?Many training and professional development opportunities were provided and include the training of nine summer students and technicians in a Computer Vision Academy, organized by the VitisGen2 powdery mildew team, and presented by CornellUniversity and USDA-ARS, Geneva scientists, technicians, and postdocs. Participants learned grapevine biology, growth and development, pathology, and remote hands-on image annotation for neural network analysis of a diverse array of traits. Their efforts resulted in an impressive database of annotated images for the project and insight into which traits are more amenable to computer vision analysis, and improved methods for vineyard imaging and analysis. Two postdoctoral researchers at UC, Davis, and Cornell were engaged to sustain trait economic evaluations and two Graduate Research Assistants at WSU and UCD were recruited to assist with data assembly and analysis, and general survey development. They were exposed to allaspects of research methods and provided with opportunities to acquire and enhance their research skills. A graduate student and another Post-doctoral Associate (Dr. Noam Reshef) at Cornell working on fruit quality analyses enhanced their knowledge on experimental design related to understanding differences in metabolite accumulation and degradation between wild Vitis and V. vinifera. Dr. Reshef's work attracted the Vaadia-BARD post-doctoral grant for performing research on the regulation of malate in wild and domesticated grape species. The award is sponsored by the USA-Israel Binational Agricultural Research and Development (BARD) fund. Eleven undergraduate students were trained on berry sampling and analytical approaches at the fruit quality laboratory at Cornell. At NDSU, three graduate students and a Post-doc were trained on leaf material collection for DNA extraction, field phenotyping for disease (powdery mildew, downy mildew), phenology (budbreak, bloom, veraison, wood acclimation), and fruit composition. One graduate student at SDSU was trained in phenotyping and genetic analysis. At Parlier, two undergraduate students were trained on field evaluations for fruit quality, DNA extractions, and other rudimentary aspects of plant breeding. and a graduate student gained additional training on QTL mapping, especially on fruit quality traits. Three scientists from the project participated at the International Table Grape Symposium, Santiago, Chile in February 2020. Cornell and USDA-Geneva scientists participants attended the Cornell Recent Advances in Viticulture and Enology meeting in Ithaca, New York, in fall 2019. Over 51 scientists took part in a QTL pipeline training including quality control assessment organized remotely using a slack channel in January 2020. How have the results been disseminated to communities of interest?Various channels were used to communicate the research efforts and findings from the project. Conference and Summer tour events involving VG2 scientists were organized at NDSU for grape growers and enologists. The North Dakota Grape and Wine Association Annual Conference, Mandan, ND, held in February 2020 had ~60 attendees, and the use of genetic maps for localization of traits of interest, and marker-assisted selection for disease resistance were highlighted. Due to COVID-19 regulations, the North Dakota Grape and Winery Association Summer Farm Tour: Grapevine Breeding Research Update. Agriculture Experiment Station, NDSU, Fargo, ND. Sept. 2020 (Attendees: ~55) took place online. The use of genetic maps for the localization of traits of interest and marker-assisted selection for disease resistance were presented. Attendees were also shown the experimental vineyards containing individuals selected with rhAmpSeq markers and observed the variation in fruit ripeness. Attendees were visually shown relative PM resistance of individuals from a target PM resistant cross (Rip.711 x NY 10.0934.01) containing RUN1 allele, compared to individuals with no known PM resistance. Results from VitisGen2 scientists were featured in trade publications (two articles), Wine Business Monthly (circulation 14,000), American Vineyard magazine (circulation 7,700), Good Fruit Grower, and Western Farm Press. Media Services at Cornell highlighted the rhAmpSeq DNA marker platform in a Cornell Chronicle article. We use social media, webinars, and podcasts to reach our national and international audience. Our scientists - Dr. Clark and Dr. Reisch were recently featured on podcasts: https://www.organicwinepodcast.com/ and https://www.vineyardteam.org/resources/resource-library/pest-management.php?id=861, respectively, where they highlighted the work of VitisGen2. The VitisGen2 website (https://www.vitisgen2.org/) is broadly visited by both public and private grape community and the project's Twitter account (@VitisGen) is constantly used to communicate scientific news to the public and scientist alike. What do you plan to do during the next reporting period to accomplish the goals?Efforts will be intensified on the current genome sequencing project. Optical maps of V. piasezkii will be used to improve sequence contiguity. Structural and functional gene annotation of the three genome assemblies will be performed to serve as a reference for the full sibling transcriptomic analysis. Additional parental genomes will be sequenced: V. cinerea B9 (Ren2), Horizon (Ren3), Y315-43-04 (Ren1), and V. rupestris B38. A major goal is to focus on the biology of the key traits of powdery mildew and fruit quality. We plan to incorporate the rhAmpSeq data into a practical haplotype graph structure used by other breeding programs in the development of genomic selection tools. The plan for implementing this future aim is to partner with the Breeding Insight project to genotype the existing USDA germplasm. Incorporating this genome-wide data for the complete repository system will allow us to fully leverage rhAmpSeq technology to identify key germplasm for important traits. We will partner with Breeding Insight to make the newly developed user-friendly software for MAS widely available for other specialty crops, filling a void for crops where single SNPs are not useful markers for selection. Resistance gene stacks (vines with multiple resistance genes) will be distributed to the clean plant network for public distribution. Phenotypic effects and downstream RNA pathways in single-gene lines for stacks of known resistance loci as well as how those pathways are affected in stacked lines will be investigated. For unknown resistance loci, phenotypic data for genetic mapping/QTL discovery will be generated and effort will be put in place to phenotype recombinants to narrow resistance loci, improve markers, and identify candidate genes. The project will encourage the widespread distribution of Blackbird imaging robots to scientists at other institutions and locations while we continue to support central screening at Cornell. Immature leaves from at least the V. amurensis x Valley Pearl population will be provided for PM phenotyping in 2021 from USDA-Parlier. Also, we will develop a rhAmpSeq marker set for Erysiphe necator population genetics and fungicide resistance screening. For RNAseq analysis, we plan to expand our isolation, sequencing, and analysis efforts to explore differences in gene expression between stacked and unstacked resistant grapevine genotypes. This is to better understand how resistance genes network in stacked backgrounds and potentially identify optimal stacking designs to increase the resilience of disease resistance. The project will continue to implement marker-assisted breeding with additional loci on mapping families using recently obtained data. New genetic maps for populations for fruit quality and Ren10 PM resistance segregation will be pursued in Minnesota. At USDA, Parlier, more backcrosses will be performed with a variety of different parents to increase the genetic base of our stacked PM resistant hybrids. Fruit (berries) from the color population will be photographed to provide records of skin color profile of individual vines. At South Dakota, an F2 population will be developed from selected individuals in the F1 population of white riparia x Alicante Bouschet. Seed from 2020 (multiple breeding programs) will be germinated; rhAmpSeq will be employed for marker-assisted selection, and questionable marker-trait associations will be validated. Seedlings are now fruiting in groups that were selected for mono- and di-glucoside pigmentation based on AmpSeq markers. These selections will be validated in 2021 based on fruit samples. Mapping populations from different breeding programs will be processed and analyzed for fruit chemistry. Also, volatiles will be measured by GC-MS on different populations from breeders. Efforts to understand the malic acid degradation difference between wild and cultivated varieties are also planned for the coming year. Metabolomic and transcriptomic data from the time-resolved sampling of Vinifera, Riparia, and Cinerea genotypes will be integrated to detect differentially expressed genes and accumulated metabolites that are associated with the dissimilation of malate in Vinifera berries and lack of in Riparia and Cinerea genotypes. Metabolite analysis of samples stored under normoxia and hypoxia will be completed to test the hypothesis of an association between a differential response to hypoxia and malate degradation. We will continue testing the previously published markers and develop new KASP markers for transferability and accuracy at predicting cluster architecture traits. Fruit texture data will be evaluated for population 11-3527 to detect putative QTL. We will finalize the analysis of the data collected via the consumer survey and distribute the outcome of the survey on the economics of new varieties for table grapes to both the general and the scientific communities. Extension outputs will focus on the central role of mapping populations, the genetics of sex determination in grapes, etc. We will equally assess the economic impact and the effect of new variety releases in providing an alternative for table and wine grapes.

Impacts
What was accomplished under these goals? This year, we completed and validated the development of the core rhAmpSeq set with >2000 amplicons, effective for genetic map construction and marker-assisted-selection (MAS) in Vitis. The flower sex marker reported last year was folded into the core rhAmpSeq panel and is now available for screening seedlings for sex alleles - M, f, or H. We discovered 2 different H (H1 and H2) alleles in the cultivated grapevine gene pool. Over 32,544 samples across 19 programs - public and private, domestic and international, etc., were processed using the new genotyping platform. A low-cost single marker KASP platform was explored to support QTL mapping. Validated markers were used to support MAS in several breeding programs. A user-friendly software for MAS quality control and statistical interpretation was developed for breeders to use in selection. Also, the AmpSeq marker platform developed in VitisGen1 was used in the genetic studies and monitoring of PM Erysiphe necator fungus. Reference genome assemblies to promote PM resistance and gene-stacking were pursued. High-throughput and recent statistical approaches were adopted for efficient laboratory and field assessment of traits. A fee-for-service rhAmpSeq genotyping option was opened to the public and private breeders not funded by the project this year. The 'Blackbird' imaging robots for PM and DM assessments are now commercially available for purchase. We continued to pursue knowledge extension to other scientists, grape growers, and the public using various channels including but not limited to publications, conferences, and the project website (vitisgen2.org). Objective 1: Technological Innovations Driven by Trait Economics: The new marker platforms were used in MAS to screen >6000 seedlings. Target crosses to generate new populations that combine disease-resistance with high fruit quality wine, table or natural dry-on-vine raisin grape attributes were made across different locations. Overall, traits considered for MAS include PM resistance (Run1, Ren1, Ren2, Ren3/9, Ren4, Ren10), DM resistance (Rpv1, Rpv3, Rpv10, Rpv12, and new loci from amurensis), muscat, di- vs. mono-glucosides, seedlessness, and flower sex. Mapping family-customized convolutional neural network models that improved detection accuracies and downstream QTL analyses were developed for PM (hyphae and conidia) and DM quantification. The development and validation of vineyard-based imaging of diseases continued on mapping populations using the Bloomfield Robotics imaging platform developed during the SCRI-Efficient Vineyard project. A low-cost QR code system attached to trellis wires was developed for vine identification. Laboratory 'Blackbird' imaging robots were upgraded to the latest mirrorless imaging technology from Nikon to improve image quality and eliminate inherent wear issues. A collaborative robotic arm for leaf disk sample processing and arraying was implemented. Over 1500 grape genotypes were phenotyped for the identification of candidate resistant genes, gene stacking, race-specificity testing, etc., of PM and DM. Over 25,000 files (3.46 TB) of microscopy and 223,000 (2.33 TB) vineyard images were generated. A 2D cluster architecture software was trained using images collected between 2016 and 2018 for cluster-related analyses. Also, we are adopting spectral measurements for a variety of traits at Cornell. In Minnesota, the use of SPME for field evaluations of floral compounds was explored. Multilocation evaluation of F2 population (180-200 genotypes, 3 reps) continued at J. Lohr vineyard, CA and Brookings, SD. Fruit samples (>820) from breeders were analyzed for quality-related attributes. Detailed pre- and post-veraison data were collected to determine the contribution of malate accumulation, degradation, and dilution to final malate concentrations on the Horizon x Illinois 547-1 family at Cornell. The analysis of 5-year malate data on ripe fruit uncovered QTL associated with malate on chromosomes 17 and 7 explaining >40% variation in malate and are stable in >3 years of the study. Genotypes differ significantly in fruit malate concentrations and the differences between haplotypes in the detected loci were up to 6.9 g/L at ripeness. Twenty genotypes bulked into 4 malate segregating groups and will be validated with metabolomic and transcriptomic analyses. Other variations (e.g. potassium), pre-and post-veraison were equally observed. Several QTL studies were carried out on several mapping families for many fruit quality, agronomic, and disease traits including tannin, malate, flowering time, veraison time, berry weight, berry size, cluster weight, leaf trichomes, foliar phylloxera resistance, Japanese beetle feeding, leaf variegation, root architecture, bud break, veraison, wood acclimation, bud freezing resistance, etc., across institutions. Gene expression and abundance as well as ethanol, acetaldehyde, CO2 production, and internal O2 levels during fruit development and ripening on vinifera, riparia, and cinerea genotypes were monitored. Results of the analysis showed that samples are primarily differentiated by phenological stage, and to a lesser extent by species. Also, the onset of ripening is associated with hypoxic conditions in both vinifera and riparia species, and vinifera berries accumulated more pyruvate and lactate, while riparia berries accumulated up to 10-fold ethanol. Wild Vitis spp. generate more CO2 per g fresh tissue than vinifera berries, a sign of a higher rate of energy-generating metabolic pathways like respiration. Objective 2: Knowledge Extension and Application: To promote the release of grapevines with various combinations of PM resistant genes and quality traits, pollen samples with different levels of PM genes were shared among breeding institutions: Run1Ren1Ren4 (USDA-Parlier to Minnesota); Ren3Ren9, and Run1Ren6Ren7 (Cornell to USDA-Parlier), etc. Also, lines with stacked PM resistant genes will be sent to Foundation Plant Services for sharing with other U.S. breeding programs. Besides, pollens with stacked sources for PM and DM resistance were received from Oliver Trapp, Geilweilerhof, Germany, and used for crosses at Cornell. Reference genomes for characterizing the biological processes associated with PM resistant loci REN1, REN2, and REN3 using BSR-seq analysis is ongoing. Single Molecule Real-Time (SMRT) bell libraries were prepared for 4 parents: V. vinifera spp. vinifera Y315-43-04 (REN1+), 2 parents of Illinois 547-1 (REN2+): V. rupestris B38 and V. cinerea B9 (REN2+), and Horizon (REN3+/REN9+). Thirty-six cDNA stranded libraries from the 2 breeding families: V. vinifera spp. vinifera Y315-43-04 (REN1+) x V. cinerea C81-227, and Horizon (REN3+/REN9+) x Illinois 547-1 (REN2+), were sequenced in single-end 150-bp mode (NextSeq500 sequencer) resulting in ~11 million high-quality reads per sample. Also, the genome assembly of V. piasezkii DVIT2027 (REN6+/REN7+) was further scaffolded using the BioNano optical maps generated last year. The target chromosome-scale assembly will serve as a reference for the characterization of the REN6 and REN7 loci. Eight F1 accessions from the breeding population 11373 (Pap et al., 2016), from V. vinifera spp. vinifera F2-35 (REN6-/REN7-) x V. piazeskii DVIT2027 (REN6+/REN7+) cross was selected for DNA sequencing: 2 REN6+/REN7-, 2 REN6-/REN7+, 2 REN6+/REN7+, and 2 REN6-/REN7- using SMRT DNA sequencing (Pacific Biosciences or Illumina DNA sequencing (DNA-seq) due to problematic DNA extraction. To better define REN1 locus, 5 V. vinifera spp. vinifera accessions possessing a REN1-like locus (Amrine et al., 2015) were sequenced using PacBio SMRT DNA sequencing. Genome assembly was performed for 4 of them: Husseine, Karadzhandal, Khalchili, and Sochal. Genome assembly of Late Vavilov is ongoing. There are genotyping efforts to ascertain new sources of PM resistance from V. doaniana 588149, V. amurensis 588631, and B37-28.

Publications

  • Type: Other Status: Accepted Year Published: 2020 Citation: R. Naegele, M. Clark, and T. Martinson (2020). Getting the Perfect Cluster Shape: Defining Table & Wine Grape Traits with DNA Markers. American Vineyard Magazine, 79:22-25. June 2020. Malcolm Media, Fresno, CA. Online at: https://cpb-us-e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2020/06/AV620ClusterShape.pdf
  • Type: Other Status: Accepted Year Published: 2019 Citation: T. Martinson, Q. Sun, C. Zou, and L. Cadle-Davidson (2019). Grape Breeders Search for Reliable DNA Markers: Why the Pinot noir PN40024 Reference Genome is Not Enough. Wine Business Monthly. December 2019. 92-97. Online at: https://cpb-us-e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2019/12/Grape-Breeders-Search-for-Reliable-Markers-WBM-December-2019.pdf
  • Type: Other Status: Awaiting Publication Year Published: 2021 Citation: T. Martinson, B. Reisch, and R. Wiepz. (2020). The central role of mapping populations in marker-assisted grape breeding: A tale of three related mapping populations at Cornell Agritech. Wine Business Monthly, Submitted July 2020. Scheduled for Feb 2021 issue.
  • Type: Other Status: Accepted Year Published: 2020 Citation: T. Martinson, A. Kovaleski, and B. Reisch. (2020) Grape mapping populations reveal genetic variation in bloom and fruit development. Blog post at www.vitisgen2.org
  • Type: Other Status: Accepted Year Published: 2020 Citation: T. Martinson and B. Reisch. (2020) The Core Grape Genome and Cheap DNA Sequencing: A New Roadmap for Grape Breeders. Appellation Cornell Issue 42, August 2020.
  • Type: Other Status: Accepted Year Published: 2019 Citation: T. Martinson and G. Sacks. (2019) Flavor Challenges of Breeding Disease-Resistant Grapes Using North American Vitis spp. Research Focus Article, Appellation Cornell Issue 39, November 2019.
  • Type: Other Status: Accepted Year Published: 2019 Citation: J. Van Zoeren, T. Martinson, C. Ledbetter, M. Clark, and B. Reisch 2019. Grape Selections from the VitisGen and VitisGen2 Projects.
  • Type: Other Status: Published Year Published: 2020 Citation: Lee Allan. 2020. Genetic Markers are the Future of Viticulture Western Farm Press, September 2020
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Sarah Thompson. 2020. Genetic marking discovery could ease plant breeders work. Cornell Chronicle, January 2020.
  • Type: Journal Articles Status: Submitted Year Published: 2020 Citation: Lior Gur, Moshe Reuveni, Yigal Cohen, Lance Cadle-Davidson, Breanne Kisselstein, Shmuel Ovadia, Omer Frenkel (2020). Population structure of Erysiphe necator on domesticated and wild vines in the Middle East shed new light on origins of the grapevine powdery mildew pathogen. Molecular Ecology.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Lance Cadle-Davidson. 2020. Computer vision quantification of foliar disease severity. Northeast Division American Phytopathological Society Annual Meeting, in Northampton, Massachusetts, March 11-13, 2020.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Cheng Zou, Avinash Karn, Bruce Reisch, Allen Nguyen, Yongming Sun, Yun Bao, Michael S. Campbell, Deanna Church, Stephen Williams, Xia Xu, Craig A Ledbetter, Sagar Patel, Anne Fennell, Jeffrey C. Glaubitz, Matthew Clark, Doreen Ware, Jason Londo, Qi Sun and Lance Cadle-Davidson (2019. Haplotyping the Vitis collinear core genome with rhAmpSeq improves marker transferability in a diverse genus. Nature Communications. https://doi.org/10.1038/s41467-019-14280-1.
  • Type: Journal Articles Status: Submitted Year Published: 2020 Citation: Lu Yin, Avinash Karn, Cheng Zou, Lance Cadle-Davidson, Anna Underhill, Paul Atkins, Dan Voytas, Erin Treiber, Matthew Clark (2020) Genetic mapping and fine mapping of leaf trichome density in cold-hardy hybrid wine grape populations. Frontiers in Plant Science.
  • Type: Journal Articles Status: Submitted Year Published: 2020 Citation: Cheng Zou, Melanie Massonnet, Andrea Minio, Sagar Patel, Victor Llaca, Avinash Karn, Fred Gouker, Lance Cadle-Davidson, Bruce Reisch, Anne Fennell, Dario Cantu, Qi Sun, Jason Londo (2020). Multiple independent recombinations led to hermaphroditism in grapevine. PNAS (Submitted).
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Teh, S.L., Rostandy, B., Awale, M. et al. Genetic analysis of stilbenoid profiles in grapevine stems reveals a major mQTL hotspot on chromosome 18 associated with disease-resistance motifs. Hortic Res 6, 121 (2019). https://doi.org/10.1038/s41438-019-0203-x
  • Type: Books Status: Published Year Published: 2019 Citation: The Grape Genome, Cantu and Walker (Eds). Springer International Publishing. Number of Pages: XXVII, 367; DOI: 10.1007/978-3-030-18601-2.
  • Type: Other Status: Other Year Published: 2019 Citation: Cantu, D. The role of domestication, breeding, and clonal propagation on genomic diversity in grapevines. Max Planck Institute for Developmental Biology. February 12, 2019, T�bingen, Germany.
  • Type: Other Status: Other Year Published: 2019 Citation: Cantu, D. Leveraging plant and microbial genomics to study grapevine diseases. Penn State University. February 25, 2019, State College, PA.
  • Type: Other Status: Other Year Published: 2019 Citation: Cantu, D. Challenges Impacting the California Wine Industry. UC Davis Chancellors Board of Advisors. April 19 2019. Napa, CA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Cantu, D. The wild side of grapevine genomics. Plant and Animal Genome XXVIII Conference. January 12, 2020, San Diego, CA.
  • Type: Other Status: Other Year Published: 2020 Citation: Anne Fennell. Cold Genetics. North Dakota Grape and Wine Association Conference, February 29, 2020, Mandan, ND.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2020 Citation: Roberto Villegas-Diaz, Dilmini Alahakoon, Jason Londo, Anne Fennell. MetaPipe: A High-Performance Computing pipeline for QTL mapping of large metabolic datasets. XXII Simposio Internacional de M�todos Matem�ticos Aplicados a las Ciencias. February 25-28, 2020, Costa Rica.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Zou C, Gouker F, Karn A, Fennell A, Cantu D, Xu X, Clark M, Reisch B, Sun Q, Cadle-Davidson L, Londo J. Molecular Characterization of the Sex Loci in Wild and Domesticated Grapes. Plant and Animal Genome Meeting. San Diego, CA. January 12, 2020.
  • Type: Other Status: Other Year Published: 2020 Citation: Roberto Villegas-Diaz. Parallel computing using Rmpi. XXII Simposio Internacional de M�todos Matem�ticos Aplicados a las Ciencias February 25-28, 2020, Costa Rica. Git hub repository: https://github.com/villegar/xxii-simmac.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Maher, M. F., Nasti, R. A., Vollbrecht, M., Starker, C. G., Clark, M. D., & Voytas, D. F. (2019). Plant gene editing through de novo induction of meristems. Nature Biotech. doi: https://doi.org/10.1038/s41587-019-0337-2
  • Type: Journal Articles Status: Accepted Year Published: 2020 Citation: Underhill, A. N., Clark, M. D., & Hirsch, C. D. Image-based phenotyping identifies QTL for cluster compactness in grape. J ASHS. [Accepted 6/14/20].
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Underhill, A. N., Clark, M. D., & Hirsch, C. D. (2020). Evaluating and mapping grape color using image-based phenotyping. Plant Phenomics: 8086309. https://doi.org/10.34133/2020/8086309.
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Yin, L., Clark, M., Burkness, E., & Hutchison, W. (2019). Grape Phylloxera, Daktulosphaira vitifoliae Fitch (Hemiptera: Phylloxeridae), on Cold-hardy Hybrid Wine Grapes (Vitis spp.): A Review of Pest Biology, Damage, and Management Practices. J. Integrated P Genetic understanding of resistance to foliar phylloxera, Daktulosphaira vitifoliae Fitch, in cold-hardy hybrid grapesest Management. doi: https://doi.org/10.1093/jipm/pmz011.
  • Type: Theses/Dissertations Status: Submitted Year Published: 2020 Citation: Olson, J. Genetic analysis and characterization of variegation in hybrid grape populations (Vitis spp.)
  • Type: Theses/Dissertations Status: Submitted Year Published: 2020 Citation: Yin, L. Genetic understanding of resistance to foliar phylloxera, Daktulosphaira vitifoliae Fitch, in cold-hardy hybrid grapes
  • Type: Theses/Dissertations Status: Accepted Year Published: 2020 Citation: Alahakoon, Dilmini. Exploring Phenotypic Diversity and Quantitative Trait Loci Mapping for Root Architecture, Freezing Tolerance, Chilling Fulfillment, and Photoperiod Traits in Grapevine Populations.
  • Type: Journal Articles Status: Submitted Year Published: 2020 Citation: Yin, L., Karn, A., Zhou, C., Cadle-Davidson, & Clark, M.D. Fine mapping of a foliar phylloxera resistance locus revealed candidate genes in a large population of hybrid grape. [Submitted to Frontiers in Plant Science]
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2020 Citation: Moreira, L., Wannemuehler, S.D., Treiber, E., Suresh, J., Brockman, S., Clark, M.D., & Hegeman, A. Sensory and metabolomic analyses link attributes of flavor and aroma in North American cold-hardy grapes. American Society for Horticultural Science, Virtual Conference. (August 10-13, 2020)
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2020 Citation: Clark, M.D., Treiber, E., Karn, A., Zou, C., Cadle-Davidson, L., & Reisch, B. Seedling selection with AmpSeq Enriches Populations for Seedlessness, Muscat Aroma, and Powdery Mildew Resistance. 9th International Table Grape Symposium, Santiago, Chile. (February 17-21, 2020).
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2020 Citation: Moreira, L., & Clark, M.D.�Characterization of flavor and aroma compounds in Minnesota cold-hardy grapes. 9th International Table Grape Symposium, Santiago, Chile. (February 17-21, 2020).
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2020 Citation: Treiber, E., Moreira, L., & Clark, M. D. Cold-hardy table grapes in Minnesota. 9th International Table Grape Symposium, Santiago, Chile. (February 17-21, 2020).
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Elizabeth M. Demmings, Brigette R. Williams, Cheng-Ruei Lee, Paola Barba, Shanshan. Yang, Chin-Feng Hwang, Bruce I. Reisch, Daniel H. Chitwood and Jason P. Londo (2019). Quantitative Trait Locus Analysis of Leaf Morphology Indicates Conserved Shape Loci in Grapevine. Frontiers in Plant Science 10: 1373.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Chin-Feng Hwang, Surya D. Sapkota, Li-Ling Chen and Karlene Negus (2020). QTL Mapping of Botrytis Bunch Rot Resistance in a Vitis aestivalis-derived Norton-based Population. Abstract for the 2020 Show Me Grape and Wine Conference and Symposium
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Chin-Feng Hwang and Li-Ling Chen (2019). Optimizing Grape Breeding with Marker-Assisted Selection. Abstract for the 2019 International Conference on Grape Genomics and Genetic Breeding.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Barba, P., Loughner, R., Wentworth, K., Nyrop, J.P., Loeb, G.M. and Reisch, B.I. 2019. A QTL associated with leaf trichome traits has a major influence on the abundance of the predatory mite Typhlodromus pyri in a hybrid grapevine population. Horticulture Research 6, 87 https://doi.org/10.1038/s41438-019-0169-8.
  • Type: Journal Articles Status: Accepted Year Published: 2019 Citation: Fresnedo-Ram�rez, J., S. Yang, Q. Sun, A. Karn, B.I. Reisch, and L. Cadle-Davidson. 2019. Computational analysis of AmpSeq data for targeted high throughput genotyping of amplicons. Front. Plant Sci. 14 May 2019 / doi: 10.3389/fpls.2019.00599.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Demmings, E.M., B. Williams, C.R. Lee, P. Barba Burgos, S. Yang, C.F. Hwang, B.I. Reisch, D.H. Chitwood, and J.P. Londo. 2019. QTL analysis of leaf morphology indicates conserved shape loci in grapevine. Front. Plant Sci. doi: 10.3389/fpls.2019.01373
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Zou, C., A. Karn, B. Reisch, A. Nguyen, Y. Sun, Y. Bao, M.S. Campbell, D. Church, S. Williams, X. Xu, C.A. Ledbetter, S. Patel, A. Fennell, J. Glaubitz, M. Clark, D. Ware, J.P. Londo, Q. Sun, and L. Cadle-Davidson. 2020. Haplotyping the Vitis collinear core genome improves marker transferability in a diverse genus. Nature Communications doi: 10.1038/s41467-019-14280-1.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Sapkota, S. D. Martinez, B. Reisch, D. Gadoury, and L. Cadle-Davidson. 2019. Do resistance genes act synergistically against grapevine powdery mildew (Erysiphe necator)?. Plant Health 2019, APS Annual Meeting, 3-7 August 2019, Cleveland, OH, USA (abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Reisch, B.I. and L. Cadle-Davidson. 2019. Perspectives from the VitisGen2 project; Update from Genevas grape breeding program. 2019 North American Grape Breeders Conference. 15-16 August 2019. Missouri State University, Columbia, MO, USA. (abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Fennell, A., D. Alahakoon, A. Karn, C. Zou, Q. Sun, J. Londo, B. Reisch, L. Cadle-Davidson, and J. Luby. 2019. Genetic analysis of grapevine root system architecture. 2019 North American Grape Breeders Conference. 15-16 August 2019. Missouri State University, Columbia, MO, USA. (abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Reshef, N., E.A. Burzynski-Chang, A. Karn, J. Londo, B. Reisch, G.L. Sacks. 2019. The Real Sour Grapes: Integrated QTL Mapping & Omics to Elucidate Malic Acid Regulation Across Grapevine Species. Society for Experimental Biology Annual Meeting. September 2019. (abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2020 Citation: Reshef, N., E.A. Burzynski-Chang, A. Karn, J. Londo, B. Reisch, G.L. Sacks. 2020. The real sour grapes  elucidating malate regulation across grapevine species. 71st American Society for Enology and Viticulture National Conference. 15-18 June 2020. Portland, OR.
  • Type: Conference Papers and Presentations Status: Awaiting Publication Year Published: 2020 Citation: Reisch, B.I. 2020. Cornells grape breeding program and the VitisGen2 project. 2020. Annual Meeting of the Virginia Vineyards Association. 20 Feb. 2020. Charlottesville, VA.
  • Type: Other Status: Published Year Published: 2019 Citation: Martinson T and Sacks G. "Flavor Challenges of Breeding Disease-Resistant Grapes Using North American Vitis spp. Appellation Cornell. Issue 39, Nov 2019.
  • Type: Other Status: Published Year Published: 2020 Citation: Naegele RP, Clark M, Martinson T. Getting the perfect cluster shape: defining traits and developing DNA markers American Vineyard magazine. Vol. 29. June 2020.
  • Type: Other Status: Other Year Published: 2020 Citation: Carlos Martinez-Meija, Breanne Kisselstein and David Gadoury. 2020. Host-Microbe Interactions: a preliminary study of an obligate biotroph, Erysiphe necator, and how it is influenced by its grape host types. Poster presented at the National Summer Undergraduate Research Project (NSURP).
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Julian M. Alston, Torey Arvik, Jarrett Hart, and James T. Lapsley. 2020. Brettanomics I: The Cost of Brettanomyces in California Wine Production. Journal of Wine Economics 128. doi:10.1017/jwe.2020.20
  • Type: Journal Articles Status: Published Year Published: 2020 Citation: Jarrett Hart, and Julian M. Alston. 2020. Evolving Consumption Patterns in the U.S. Alcohol Market: Disaggregated Spatial Analysis. Journal of Wine Economics 15(1), 541. doi:10.1017/jwe.2019.14
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Hart, Jarrett, and Julian M. Alston. 2019. Under the (Ancestral) Influence: Demographic Drivers of Diverging Demands in the U.S. Alcohol Market. ARE Update 23(2): 58. University of California Giannini Foundation of Agricultural Economics.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Jarrett Hart, and Julian M. Alston. 2019. Persistent Patterns in the U.S. Alcohol Market: Looking at the Link between Demographics and Drinking. Journal of Wine Economics 14(4), 356364. doi:10.1017/jwe.2019.26
  • Type: Book Chapters Status: Published Year Published: 2019 Citation: James T. Lapsley, Julian M. Alston, and Olena Sambucci. 2019. The U.S. Wine Industry. Chapter in Adeline Alonso Ugaglia, Jean-Marie Cardebat, and Alessandro Corsi (eds) the Palgrave Handbook of Wine Industry Economics.


Progress 09/01/18 to 08/31/19

Outputs
Target Audience:Various interest groups were reached by VitisGen2 efforts this year including the scientific community, stakeholders, industry members, extension agents, students, and the public through diverse channels not limited to conferences, workshops, seminars, presentations, field days, publications, websites, and social media. VitisGen2 scientists presented at numerous conferences locally and internationally covering a range of topics such as advances in grape genotyping, genetic and genomic improvement strategies, high throughput phenotyping, disease resistance, economic aspects, etc. to the scientific community, extension agents and industry representatives. The scientific community and the general public also benefited from the numerous publications in top peer-reviewed journals, trade publications, news articles, fact sheets, field days, seminar presentations, workshop presentations, stakeholder board meetings, etc. The VitisGen2 website (www.Vitisgen2.org), and the social media twitter account were actively used to communicate project findings to both the scientific and non-scientific audience especially through the "Research in Plain English" initiative. Field days, meetings and presentations were used to engage the host communities, scientists, family farmers, end-users, students and the general public on the activities of the team and research outputs. The robot laboratory hosted scientists and the public, such as high school students interested in engineering; graduate students in plant breeding; stakeholders from the grape and biocontrol industries; USDA national program leaders; congressional representatives; and economic development experts and communicated regarding the multidisciplinary (biology, informatics, engineering) and multi-institutional research approach as well as the role of applied robotics, statistics and computation in modern agriculture. Data and protocols developed by the team were most accessible to scientists and the public. There were other forms of training offered to students at all levels from high school students to post-doctoral and visiting scholars, including visitors abroad. Online webinars from the team were used to reach scientists, grape industry members, students, etc. and are recorded/posted online for anyone to view. Survey tools were designed and used to directly engage consumers and stakeholders. Various discussions and dialogues aimed at capturing and characterizing end-user preferences that could help modify breeding goals and objectives were explored. Changes/Problems:There were no problems warranting a deviation from the proposed objectives and goals. The few recorded challenges within the reporting period were adequately resolved in keeping with the expected goals. Some issues with DNA extraction quality were identified and later traced to sampling issues. This was adequately addressed in collaboration with the DNA isolation contractor (Intertek), IDT and Cornell sequencing facilities. There were delays in the volatile phenotyping of VitisGen2 populations due to problems with the main instrument used for the analyses (LECO Pegasus GC-MS). The use of the LECO device was decommissioned and the methods were translated to a different instrument (Shimadzu 8040 GC-MS/MS). This will be used to continue volatile phenotyping in the future. Also, there was a technical challenge with the Orbitrap LC-MS based metabolomics sampling for malic acid metabolism studies. Although negative ion mode analysis was successful, positive ion mode analyses yielded irreproducible results for amino acids and several other metabolites. We are transitioning to the GC-MS platform for future metabolomic analyses. Some of our fruit quality validation populations had low vigor and vine numbers were insufficient for validating markers. Other internal delays were related to hiring which brought some delays in the delivery of a few expected outcomes. For example, the 3D imaging boxes were expected to be available in July, but are not expected until next spring due to manpower. What opportunities for training and professional development has the project provided?The project provided numerous training and professional development opportunities for various partners at different levels within this reporting year. The VitisGen2 scientists participated in notable conferences including the American Society for Enology and Viticulture-Eastern Section (ASEV-ES) and the 2019 North American Grape Breeders conferences where they participated in various scientific deliberations and training on high throughput phenotyping of grapevines. Members were also trained on Mixed modeling using R at the American Phytopathological Society (APS) annual meeting. Other training included: Introduction to Linux; bioinformatics in Linux; RNA-Seq data analysis; genome sequence data analysis; genome annotation and sequence-based gene function prediction at the Cornell University Institute of Biotechnology; Linear and multiple regression; Intermediate data analysis in R at the Cornell University Statistical Consulting Unit; and operator training on Universal Robots at Cornell AgriTech. At ARS-Parlier, a technician was trained to distinguish between seeded and stenospermic fruits and exposed to the diversity of seed trace sizes present in stenospermic vines and how to differentiate them from seeded accessions. Undergraduate and high school students were trained on the different aspects of in-ovulo embryo culture, aseptic technique, medium preparation, fruit disinfestation, berry sampling and analytical approaches, ovule placement, and embryo identification. Others were trained in data assembly, and analysis, as well as survey development and design. A total of 13 undergraduate and 2 high school students received training from VitisGen2 this current year. Two graduate students and a post-doctoral associate at Sacks' lab in NY-Cornell received training on experimental design related to understanding differences in metabolite accumulation and degradation between wild Vitis and V. vinifera. Other graduate students were trained in field evaluations of fruit quality, DNA extraction, and other aspects of plant breeding. Some Ph.D. students were added to the project at the University of Minnesota, South Dakota State University and Missouri State University and received training on genetic mapping studies. Visiting scholars and post-doctoral researchers equally received new skills and were exposed to multidisciplinary research opportunities for grape improvement. How have the results been disseminated to communities of interest?VitisGen2 utilized various channels to disseminate research findings and discoveries to various audiences including students, the scientific community, industry, stakeholders, extension agents, and the general public both locally and abroad. The VitisGen2 work was featured in notable magazines such as the Wine Enthusiast magazine, e.g. "How Computers are Being Used to Hack Grapes" by Stacy Brooks on April 2, 2019 (https://www.winemag.com/2019/04/02/computers-hack-wine-grapes/). Over 100 attendees were present at the VitisGen2 research presentations at the Minnesota Grape Growers Association Cold Climate Conference of the University of Minnesota Grape Breeding and Enology project. Our scientists were featured at the quarterly board meeting updates of the Minnesota grape industry and at the semi-annual board meeting of the National Grape Research Alliance. Four grower workshops in association with an NC-SARE partnership grant on grape growing focused on disease resistance management practices and measuring fruit quality traits to improve harvest, conducted at MN. An open house was held at the Horticultural Research Center, Excelsior, MN in Sept 2018 where presentations and posters on grape tasting for potential new varieties were presented to the public, industry, and stakeholders numbering up to 75. VitisGen2 objectives and results were frequently disseminated in lectures requested by local educators at USDA-Parlier. Information on durable PM resistance breeding was made available locally and internationally to many including 8 Ukrainian Ag-Econ. teachers on a Foreign Agricultural Service teacher's exchange program with California State University, Fresno; 19 summer interns employed at a table grape nursery; and 8 Serbian wine industry professionals on a FAS training program with CSUF. A field day was organized at Cornell-Geneva for scientists at Ithaca and other localities. This field day included the Breeding insight project team during August 2019. One of the papers on the grape industry was published as a chapter in a book, published by the Giannini Foundation, targeting diverse audiences and available to the general public. The powdery mildew phenotyping laboratory (also known as the Robot Lab) was a frequent site of outreach events focused on communicating the importance of multidisciplinary (biology, informatics, engineering) and multi-institutional research, with about 20 tours provided to scientists and the public, including high school students interested in engineering; graduate students in plant breeding; stakeholders from grape and biocontrol industries; USDA national program leaders; congressional representatives; and economic development experts. VitisGen2 articles and webinars highlighted the need for new varieties to address genetic weaknesses in current varieties, and how advances in DNA technology have made it possible to map and discover traits for disease resistance, insect resistance, cluster architecture, color, and sex determination. Several such articles in general media outlets (Forbes.com, NPR, Smithsonian) featured contributions from project partners. The rhAmpSeq platform developed by VitisGen2 has received more general interest from geneticists and breeders of other crops. What do you plan to do during the next reporting period to accomplish the goals?For the future, one major goal is to incorporate the rhAmpSeq data into the practical haplotype graph structure used by other breeding programs in the development of genomic selection tools. The key to implementing this future aim is to partner with the Breeding Insight project to genotype all existing USDA Vitis germplasm. Incorporating this genome-wide data for the complete repository system will allow us to fully leverage rhAmpSeq technology to identify key germplasm for important traits. The team will capitalize on the advances of rhAmpSeq markers as well as the ongoing work in RNAseq and genome sequencing. Optical map data of V. piasezkii will be used to improve sequence contiguity. Structural and functional gene annotation of three existing genome assemblies will be performed to serve as a reference for the full sibling transcriptomic analysis. Additional parental genomes will be sequenced. Also, there will be an effort to construct a Chambourcin high-resolution linkage map with both SSR and rhAmpSeq-based SNP markers. We will continue to focus on phenotypic evaluations and understanding the biology of key traits especially powdery mildew and fruit quality. An effort will be made to encourage the widespread distribution of Blackbird imaging robots to scientists at other institutions and locations as well as transferring resistance gene stacks (vines with multiple resistance genes) to repositories and/or the national clean plant network (NCPN) for public distribution. Attention will be given to stacks of known resistance loci to investigate phenotypic effects, downstream RNA pathways in single-gene lines, and how those pathways are affected in stacked lines. On the other hand, for unknown resistance loci, we will generate phenotypic data for genetic mapping/QTL discovery. There are plans to expand RNAseq isolation, sequencing, and analysis efforts to explore differences in gene expression between stacked and unstacked resistant grapevine genotypes. This is to better understand how resistance genes network, or interact, in stacked backgrounds and potentially to identify optimal stacking designs to increase the durability of disease resistance. The analyses of the fruit chemistry of samples submitted by our VitisGen2 collaborators will continue at the CFQC. This will include two new populations harvested in 2019, from NY-Cornell (NY84.0101.03 x V.amurensis), and NY-USDA (V. doaniana 588149 x Chardonnay). Also, 2019 tannin analysis submitted from Missouri will be carried to further investigate tannin QTL in hybrid grapes. We will also complete volatile analyses, which were delayed due to challenges with the GC-MS instrument. The time-resolved studies of malic acid and minerals in vinifera and wild Vitis will continue. RNA libraries will be created and RNAseq analysis conducted on previously collected samples to evaluate if phenotypes can be explained by differences in gene expression. We will also complete metabolomic analyses on 2018 samples. We will phenotype the Horizon X Illinois population at pre- and post-veraison stages for an additional year (2019), to verify QTL identified for malate in 2018. In addition, we will verify the correct selection of segregating groups and target them for further metabolomic and transcriptomic analysis. Based on these analyses and subsequent results, we plan to highlight candidate genes that are regulating malate levels in the grape. Efforts to understand malic acid degradation difference between wild and cultivated varieties are also planned for year 3 of the grant. The use of x-ray in addition to 3D RGB imaging will be deployed to evaluate grape clusters at USDA-Parlier. We have also identified new validation populations from more established populations to offset the small population numbers in our validation populations. The initiated consumer survey will be completed and analyzed and research outcomes pertaining to new varieties, as well as research findings, will be disseminated to the general public and the necessary academic audiences. The project will continue to expand on current outreach programs using webinars, spotlight profiles, "Research In Plain English" summaries, trade articles and journals in communicating research outputs. Also, field days will be used to strengthen collaborations with growers and industries. A field day in the Central Valley of California in association with Keith Striegler, EJ Gallo is expected in 2020.

Impacts
What was accomplished under these goals? Objective 1: Technological Innovations Driven by Trait Economics To support genome-guided improvement strategies in grape, the ongoing development and testing of the core rhAmpSeq probe set for Vitis continued this year. Twelve genomes of Vitis were sequenced and assembled, then used to identify core conserved sequences, resulting in a 2000 amplicon set, with over 95% of the markers returning useful data across multiple families. An additional 55 markers were added to the 2000 core genome amplicons for marker-assisted selection. Multiplexing of these amplicons led to a reduction in cost per sample from ~$20 to under $10 USD. We are integrating the rhAmpSeq and the previous genotype-by-sequencing (GBS) data from VitisGen1 into a single reference set of genomic coordinates. This year, 41 QTL were mapped across 9 mapping families, new linkage maps constructed, and marker-assisted seedling selection for disease resistance (RUN, REN and RPV loci), muscat flavor, seedlessness and seed trace size were carried out with the previously validated markers. A total of 165 markers (multiple markers per trait) were assessed across 6,425 seedlings. All markers are being converted for use with rhAmpSeq techniques. Rapid phenotyping of economically important traits remains a high priority. A robotic arm was developed for leaf disk sampling and arraying preceding mildew inoculation, capable of ~2800 samples per day. Four Blackbird imaging robots, commercial versions of the prototype developed earlier in the project, were installed at NY-USDA (processes ~10,000 samples per day). Two new convolutional neural network algorithms were developed for rapid data analyses and quantification of grape powdery mildew (PM) hyphae and conidia. A total of 3413 genotypes were assayed for PM or downy mildew (DM) reactions to support the ongoing genetic mapping, marker validation, gene stacking, fine mapping, and race-specificity testing. Leaf tissue samples were received from USDA-Parlier, NY-USDA, MO, SDSU, and NY-Cornell. Ca. 2000 vines were screened to identify recombinants between the REN3 and REN9 PM resistance loci. Initial testing of a field-based imaging system for field-based phenotyping using an ATV-mounted camera began in 2019. A total of 150,000 and 500,000 images were generated from the Blackbird and vineyard imaging, respectively. New collaborations to develop an image-based analysis for seed trace size and 3D scans of fruit architecture were also initiated. The Cornell Food Quality/Wine Center (CFQC) and Donald Danforth Center in Missouri evaluated samples for quality components. A total of 357 grape samples were processed for pH, Brix, berry weight, and titratable acidity; 403 (malic acid concentration); 315 (acetone-extracted tannin); 179 (ethanol-extracted tannin) at the CFQC; and 165 for metals at the Danforth Center. Mapping populations were from MN, MO, and NY-Cornell, and NY-USDA. In addition, metabolomic analyses were performed on wild V. cinerea, V. riparia accessions and V. vinifera (Finger Lakes, NY) sampled in 2017 and 2018. Marker-assisted selection of progenies segregating for malate was carried out and samples used for bulked segregant analysis. Transcriptomic studies were initiated to further understand the degradation pathway of the fruit quality complex in Vitis. At the Table/Raisin Quality Center, ARS-Parlier, samples were evaluated for ripening time, time to full color, cluster imaging, berry texture, etc., and genotyped with rhAmpSeq. Previously published molecular markers associated with fruit architecture were tested and found to be nontransferable to table grape populations. A 20 fold variation in acetone-extractable tannin was observed from a mapping population at MO. Another family differed in fruit malate concentrations at both pre- and post-veraison stages. Two preliminary QTLs were detected with 2017 data, although with low statistical significance due to small numbers of fruiting vines. The analysis of 2018 samples is underway. Samples of 20 genotypes were bulked into 4 segregating groups of progenies exhibiting contrasting pre- and post- veraison malate trends. Further metabolomic and transcriptomic studies were initiated in the 4 groups. Time-resolved sampling and analysis of vinifera and wild Vitis spp. grapes showed that wild Vitis accumulate malate at higher levels pre-veraison, and dissimilate less malate post-veraison (ripening). Specific measurements of malate levels in mesocarp tissue showed that these differences are not the results of a higher skin/pulp ratio in wild Vitis, but rather physiological differences in the mesocarp tissue. Field evaluations for PM resistance were carried out in mapping populations at MO, NY-Cornell, NY-USDA, and MN. Other traits evaluated include bloom date, root architecture, DM, leaf trichomes, botrytis bunch rot, time of bud break, etc. New sources of PM resistance from wild species (V. riparia, V. cinerea, V. vulpina, and V. amurensis) were explored. Notable vines demonstrating PM resistance from a V. amurensis x Valley Pearl population were backcrossed with high-quality table grapes and natural dry-on-vine raisins at USDA-Parlier. The use of RNAseq analysis to understand the functional aspects of powdery mildew resistance was initiated. Pooled samples of resistant and susceptible material will allow the differentiation of gene expression of candidate genes and downstream pathways. The inheritance of leaf variegation is being studied using populations from MN and NY. Additionally, a fine-mapping population (n=1000) at MN was phenotyped for leaf phylloxera resistance, stomatal conductance, and photosynthesis. Genetic analysis of phylloxera resistance resulted in the haplotype marker identification of a locus which agreed with a previously reported QTL in the literature. To study the effect of genotype x environment interactions on important grape traits, a V. riparia x Seyval population is being evaluated at two locations in three replicates. Previously published flower sex markers do not work well in mapping families that utilize wild grape germplasm. The use of RhAmpSeq markers combined with genome sequencing of diverse germplasm was a powerful tool for complex sex locus studies. We found strong evidence for two specific candidate genes leading to the expression of male and female flowers. Results are being validated but currently support the identification of all three flower types at the seedling stage. To evaluate consumer preferences and responses to new products, the adoption study of grape varieties was initiated. Online consumer surveys to evaluate consumer attitudes towards specific traits/varieties, as well as consumer response to technologies such as gene-editing, was launched and is expected to be administered to over 2,000 participants. A new collaboration with postharvest researchers to link fruit texture values with consumer taste panels was also initiated. Objective 2: Knowledge Extension and Application Knowledge extension and application are documented here and in later sections. These included targeting a broad audience of other scientists, growers, industry leaders, and the general public through the project website (Vitisgen2.org), where we post project news, accomplishments, technical advances, and goals of the VitisGen2 project. In addition, webinars, trade publication articles, twitter feed and non-technical summaries of VitisGen2 journal articles are detailed below. Across the breeding programs, the use of marker-assisted selection was widely adopted with validated markers for PM, DM, leaf phylloxera resistance, muscat flavor, seedlessness and seed trace size. There are ongoing efforts to explore more PM resistance genes from the wild in addition to gene stacking efforts for multiple combinations of RUN1, REN1, REN2, REN3, REN4, REN6, REN7, and REN10 PM resistance.

Publications

  • Type: Theses/Dissertations Status: Published Year Published: 2019 Citation: Underhill, A. (2019) Using high-throughput phenotyping to investigate the genetic bases of quantitative traits in hybrid wine grapes (Vitis spp.). University of Minnesota Masters Thesis
  • Type: Other Status: Submitted Year Published: 2019 Citation: Martinson, T., Q. Sun, C. Zou, and L. Cadle-Davidson. (2019). Grape breeders search for reliable DNA markers: Why the Pinot Noir PN40024 reference genome is not enough. Submitted to Wine Business Monthly, July 2019.
  • Type: Other Status: Published Year Published: 2019 Citation: Moyer, M. M., T. Martinson and L. Cadle-Davidson (2019). Disease-resistant varieties are on the way: Can we ensure they last? Wine Business Monthly, June 2019. 76-83. Online at: https://cpb-us-e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2019/08/Moyer-martinson-Disease-Resistant-Varieties-on-the-Way.pdf.
  • Type: Other Status: Published Year Published: 2018 Citation: Martinson, T. and L. Cadle-Davidson (2018). The Phenotyping Bottleneck: How grape breeders link desired traits to DNA markers. Wines and Vines, December 2018. 142-145. Online at: https://cpb-us-e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2019/01/The-Phenotying-Bottleneck-_-Wines-and-Vines-Dec-2018-1w6gc3o.pdf.
  • Type: Other Status: Published Year Published: 2019 Citation: Martinson, T. and C. Ledbetter and R. Naegele (2019). Marker-Assisted Selection Makes Efficient Table Grape Breeding. American Vineyard, March 2019. 10-12. Online at: https://cpb-us-e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2019/03/AV201903-Breeding-2a0n2qj.pdf.
  • Type: Other Status: Published Year Published: 2019 Citation: Michelle M. Moyer, Timothy Martinson and Lance Cadle-Davidson. 2019. Disease-resistant Varieties are on the Way: Can we ensure they last? Wine Business Monthly, July 2019: pp. 76-83.
  • Type: Websites Status: Published Year Published: 2019 Citation: VitisGen2 website (http://www.vitisgen2.org/): New content includes 4 popular press articles, 4 Research in Plain English articles, 6 Staff Spotlight profiles, 2 webinars, and 4 news articles/blog posts. Statistics: 6,238 visits (2638 unique visitors), 13,552 page views, 42% of visits were new visitors.
  • Type: Websites Status: Published Year Published: 2019 Citation: @VitisGen (https://twitter.com/VitisGen)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Chin-Feng Hwang, Li-Ling Chen, Bryce Bentley and Sadie Land (2019). Optimization of Chambourcin Grape Breeding Using Molecular Genetic Approaches. Abstract for the 2019 Show Me Grape and Wine Conference and Symposium.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Chin-Feng Hwang, Li-Ling Chen, Sadie Land and Bryce Bentley (2019). Optimization of Chambourcin Grape Breeding Using Marker-Assisted Selection. Abstract for the 44th Annual Conference of the American Society for Enology and Viticulture (ASEV)-Eastern Section.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Sapkota, S. D. Martinez, B. Reisch, D. Gadoury, and L. Cadle-Davidson. 2019. Combined effects of stacking multiple resistance genes upon grapevine powdery mildew (Erysiphe necator). International Plant and Animal Genome Conference XXVII. 12-16 January 2019. San Diego, California, USA. (abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Sapkota, S. D. Martinez, B. Reisch, D. Gadoury, and L. Cadle-Davidson. 2019. Do resistance genes act synergistically against grapevine powdery mildew (Erysiphe necator)?. Plant Health 2019, APS Annual Meeting, 3-7 August 2019, Cleveland, OH, USA (abstr.)
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Gavin L. Sacks. Not to everyones taste  an update on research in the Sacks Lab. Cornell Recent Advances in Viticulture and Enology (CRAVE). Ithaca, NY. Dec 11, 2018.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Noam Reshef, Burzynski-Chang EA, Avinash Karn, Jason Londo, Bruce Reisch, Gavin L. Sacks. Elucidating the molecular regulation of grape sourness by integrating QTL mapping with RNAseq and metabolomics of non-domesticated grapevine species. Society of Experimental Biology Meeting. Seville, Spain.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Terry L Bates, Madeleine Y Bee, Xuefei Kuang, and Gavin L Sacks GL. Simultaneous Measurement of Key Odorants at their Sensory Thresholds in Juice Grapes. 67th Annual Conference of the American Society for Mass Spectrometry. Atlanta, GA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Zou, C. High-fidelity rhAmpSeq" primers targeting the core genome enables shared exploration of highly diverse and heterozygous species� at Grape Genome Initiative workshop, International Plant & Animal Genome XXVII,�January 12-16, 2019� - San Diego, CA, USA
  • Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Belton, J., Kisselstein, B. M., Gadoury, D. M. Mating Type Distribution and Formation of Chasmothecia on Grapevine by Erysiphe necator in the Northeastern United States. Summer Research Scholars Poster Session in Geneva, NY. July 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Cheng Zou, Avi Karn, Qi Sun, Lance Cadle-Davidson, et al. High-fidelity rhAmpSeq" primers targeting the core genome enables shared exploration of highly diverse and heterozygous species. Grape Genome Initiative workshop, International Plant & Animal Genome XXVII, January 12-16, 2019 - San Diego, CA, USA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Cheng Zou, Lance Cadle-Davidson, Qi Sun, Avi Karn, et al. rhAmpSeq" 2000-Plex Amplicon Sequencing Panel Targeting Core Genome in Vitis vinifera shows Elevated Marker Transferability, Increased Specificity and Applicability to Diverse Organisms. Integrated DNA Technologies workshop, International Plant & Animal Genome XXVII, January 12-16, 2019 - San Diego, CA, USA.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Dani Martinez, Andrew Bierman, Timothy Plummer, Lance Cadle-Davidson, David Gadoury, Surya Sapkota. Blackbird: A novel high-throughput laboratory phenotyping system to quantify incidence and severity of powdery mildews. Poster at Plant Health 2019 in Cleveland, OH, August 3-7, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Dani Martinez, Surya Sapkota, Andrew Bierman, Timothy Plummer, David Gadoury, Mark Rea, Lance Cadle-Davidson. 2019. Blackbird: A next-gen imaging system for high-throughput laboratory phenotyping. Talk at Phenome 2019 in Tucson, AZ, February 6-9, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Kisselstein, B.M., Cadle-Davidson, L., and Gadoury, D.M. Genetic diversity of Erysiphe necator populations in a center of diversity: a case study of select vineyards in New York State. American Phytopathological Society Annual Meeting in Cleveland, OH. August 3-7, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Lance Cadle-Davidson. Computer vision for high-throughput phenotyping of powdery mildew resistance, presented at Phenome in Tucson, Arizona. February 9, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Moreira, L.S., Underhill, A., and M. Clark. Postharvest evaluation of cold-hardy table grape breedling lines. XII International Conference on Grapevine Breeding and Genetics. .doi:10.17660/ActaHortic.2019.1248.15
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Surya Sapkota, Dani Martinez, David Gadoury, and Lance Cadle-Davidson. 2019. Phenotyping grapevine powdery mildew: an update from VitisGen2 phenotyping center. Talk at National Grape Breeders Conference, in Mountain Grove, MO, August 15-16, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Diering, A., Tork, D.G., Underhill, A., Freund, D.M., Clark, M.D., and A. Hegeman. HPLC-MS as a detection method for pigments, phenolics, and co regulation in a hybrid wine grape family & Using PCA for QTL mapping to optimize plant breeding. American Society for Mass Spectrometry, Atlanta, GA
  • Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Moreira, L.S. Hegeman, A.D., Brockman, S.A., Suresh, J., and M.D. Clark. Non-destructive field sampling of volatile organic compounds for metabolomics analysis of grape cluster development. ASEV-ES, Geneva, NY
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Yin, L. Cadle-Davidson, L., Sun, Q., Londo, J., Karn, A., Zou, C., Gouker, F., Fresnedo, J., and M. Clark. Recombinant identification using rhAmpSeq in a hybrid cold-hardy grape population for fine mapping a major resistance QTL to foliar phylloxera. Plant and Animal Genome Conference, San Diego, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Martinson, T. E. 2019 VitisGen2: Application of next generation technologies to accelerate grapevine cultivar development, Presentation, 2019 Eastern New York Fruit and Vegetable Conference Albany, NY, February 21, 2019. Audience = 20 grape producers and winery representatives from Eastern New York.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Karn, A, Zhou, C, Alahakoon, D, Fennell, A, Sun, Q, Londo, J, Reisch, B, Cadle-davidson, L. rhAmpSeq Core Genome Markers Improve the Coverage of a GBS-Based Genetic Map and Marker Transferability to Other Grapevine Populations. INTEGRAPE 2019 Data integration as a key step for future grapevine research, Chania, Crete, Greece Mar. 25-28, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Mcdermaid, A, Monier, B, Wang, C, Zhao, J, Miller, A, Fennell, A, Ma, Q. IRIS-EDA: An integrated RNA-Seq interpretation system for gene expression data analysis. INTEGRAPE 2019 Data integration as a key step for future grapevine research, Chania, Crete, Greece Mar. 25-28, 2019.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Sun Qi, Zou Cheng, Karn Avinash, Reisch Bruce, Nguyen Allen, Sun Yongming, Bao Yun, Campbell Michael S., Church Deanna, Williams Stephen, Smith Timothy P. L., Fennell Anne, Clark Matthew, Ware Doreen, Londo Jason, Cadle-Davidson Lance (2019).� Haplotype Markers Developed from the Vitis Core Genome and Their Applications for QTL Mapping and MAS Breeding in Hybrid Grape Populations. INTEGRAPE 2019 Data integration as a key step for future grapevine research, Chania, Crete, Greece Mar. 25-28, 2019.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Moreira, L.S. University of MinnesotaState Report. North American Grape Breeders Conference. Columbia, MO
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Noam Reshef. The real sour grapes  integrated QTL mapping with omics approaches to elucidate malic acid regulation across grapevine species. Society of Experimental Biology Meeting. Seville, Spain. July 2019.
  • Type: Conference Papers and Presentations Status: Other Year Published: 2019 Citation: Moreira. L.S. Using metabolomics analysis to characterize volatile compounds in cold-hardy hybrid grapes in Minnesota. ASEV National Conference, Napa, CA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Fennell, A. 15th Annual VESTA National Summit and Michigan Wine Consortium Conference, June 6, 2019, Traverse City, MI. Advancing the American grape industry through research.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Fennell, A. North American Grape Breeders Conference, Aug. 16, 2019, Springfield, Missouri. Genetic analysis of grapevine root system architecture.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Zou, C. rhAmpSeq" 2000-Plex Amplicon Sequencing Panel Targeting Core Genome in Vitis vinifera shows Elevated Marker Transferability, Increased Specificity and Applicability to Diverse Organisms at Integrated DNA Technologies workshop, International Plant & Animal Genome XXVII,�January 12-16, 2019� - San Diego, CA, USA
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Gavin L. Sacks. Speeding up SPMESH. SPME Approaches Applied to Mass Spectrometry Techniques Workshop at the 67th ASMS Meeting. Atlanta, GA. June 3, 2019.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2019 Citation: Bruce Reisch. North American Grape Breeders Conference, Aug. 16, 2019, Springfield, Missouri. Perspectives from VitisGen2 project; and updates from Geneva's grape breeding program.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Fresnedo-Ram�rez, J., Yang, S., Sun, Q., Karn, A., Reisch, B. I., & Cadle-Davidson, L. (2019). Computational analysis of AmpSeq data for targeted, high-throughput genotyping of amplicons. Frontiers in plant science, 10, 599.Doi:10.3389/fpls.2019.00599
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Andrew Bierman, Tim LaPlumm, Lance Cadle-Davidson, David Gadoury, Dani Martinez, Surya Sapkota, and Mark Rea. 2019. A High-Throughput Phenotyping System Using Machine Vision to Quantify Severity of Grapevine Powdery Mildew. Plant Phenomics 2019:92097272019. doi:10.34133/2019/9209727.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Barba, P., Loughner, R., Wentworth, K., Nyrop, J.P., Loeb, G.M. and Reisch, B.I. 2019. A QTL associated with leaf trichome traits has a major influence on the abundance of the predatory mite Typhlodromus pyri in a hybrid grapevine population. Horticulture Research 6, 87 https://doi.org/10.1038/s41438-019-0169-8
  • Type: Journal Articles Status: Accepted Year Published: 2019 Citation: Yin, L., Clark, M.D., Burkness, E.C., & Hutchison, W.D. (2019) Grape phylloxera (Hemiptera: Phylloxeridae), on cold-hardy hybrid wine grapes (Vitis spp.): A review of pest biology, damage, and management practices. J. Integrated Pest Management 10(1): 16 https://doi.org/10.1093/jipm/pmz011
  • Type: Journal Articles Status: Other Year Published: 2019 Citation: Avinash Karn, Cheng Zou, Dave Manns, Qi Sun, Jason Londo, Lance Cadle-Davidson, Anna Katharine Mansfield, Bruce I. Reisch, and Gavin Sacks (2019) Genetic Basis of Acylated Anthocyanins Variation in North-American Interspecific Grapevine Hybrids. Molecular Breeding (In process)
  • Type: Journal Articles Status: Other Year Published: 2019 Citation: Avinash Karn, Rachel Naegele, Siraprapa Brooks, Jonathan Fresnedo Ramirez, David Ramming, Craig Ledbetter, and Lance Cadle-Davidson (2019) Genetic analysis of powdery mildew resistance QTL from Vitis hybrid Tamiami. Phytopathology (In process)
  • Type: Journal Articles Status: Other Year Published: 2019 Citation: Cheng Zou, Avinash Karn, Bruce Reisch, Allen Nguyen, Yongming Sun, Yun Bao, Michael S. Campbell, Deanna Church, Stephen Williams, Xia Xu, Craig A Ledbetter, Sagar Patel, Anne Fennell, Jeffrey C. Glaubitz, Matthew Clark, Doreen Ware, Jason Londo, Qi Sun and Lance Cadle-Davidson (2019) A rhAmpSeq haplotype strategy targeting the collinear core genome improves marker transferability across a diverse genus. Nature (In process)
  • Type: Book Chapters Status: Accepted Year Published: 2019 Citation: Clark, M.D. 2019. Development of Cold Climate Grapes in the Upper Midwestern U.S.: The Pioneering Work of Elmer Swenson. In Plant Breeding Reviews Vol 43. Ed. I. Goldman. p. 31-59
  • Type: Book Chapters Status: Awaiting Publication Year Published: 2019 Citation: Julian M. Alston and Olena Sambucci. 2019. Grapes in the World Economy. In D. Cantu and M.A. Walker, eds, The Grape Genome, Compendum of Plant Genomes. Springer Nature Switzerland AG. In press.
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Burzynski-Chang, E.A., I. Ryona, B.I. Reisch, I. Gonda, M.R. Foolad, J.J. Giovannoni, and G.L. Sacks. 2018. HS-SPME-GC-MS Analyses of volatiles in plant populations  Quantitating compound x individual matrix effects. Molecules. 23(10):2436 https://doi.org/10.3390/molecules23102436
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Elizabeth A Burzynski-Chang, Elizabeth J Brown, Noam Reshef, Gavin L Sacks. Malate Content in Wild Vitis spp. Demonstrates a Range of Behaviors during Berry Maturation. American Journal of Enology and Viticulture, ajev. 2019.19015.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Sambucci, Olena, Julian M. Alston, Kate B. Fuller and Jayson Lusk. 2019. Pecuniary and non-Pecuniary Costs of Powdery Mildew Management: Evidence From California. American Journal of Enology and Viticulture 70(2): 177187.
  • Type: Journal Articles Status: Published Year Published: 2019 Citation: Surya Sapkota, Shanshan Yang, Li-Ling Chen, Lance Cadle-Davidson and Chin-Feng Hwang (2019). Construction of a High-Density Linkage Map and QTL Detection of Downy Mildew Resistance in Vitis aestivalis-derived Norton. Theoretical and Applied Genetics 132: 137-147.
  • Type: Journal Articles Status: Submitted Year Published: 2019 Citation: Demmings, E.M., B. Williams, C.-R. Lee, P. Barba Burgos, S. Yang, C.-F. Hwang, B.I. Reisch, D.H. Chitwood, and J.P. Londo. 2019. QTL analysis of leaf morphology indicates conserved shape loci in grapevine. Front. Plant Sci
  • Type: Book Chapters Status: Published Year Published: 2019 Citation: Julian M. Alston, James T. Lapsley, and Olena Sambucci. 2018. Grape and Wine Production in California. In P.L. Martin, R.E. Goodhue, and Brian D. Wright, eds, California Agriculture: Dimensions and Issues. Giannini Foundation of Agricultural Economics, University of California.
  • Type: Book Chapters Status: Awaiting Publication Year Published: 2019 Citation: Lance Cadle-Davidson, Jason Londo, Dani Martinez, Surya Sapkota, Ben Gutierrez. From Phenotyping to Phenomics: Present and Future Approaches in Grape Trait Analysis to Inform Grape Gene Function. Chapter in The Grape Genome, Eds. Dario Cantu and M.A. Walker.
  • Type: Theses/Dissertations Status: Published Year Published: 2019 Citation: Ebennga, D. Drosophila suzukii in wine grapes: Phenology, exclusion netting, and varietal resistance. MSc Thesis, University of Minnesota
  • Type: Theses/Dissertations Status: Published Year Published: 2019 Citation: Elizabeth Burzynski-Chang. Tools for the improvement of flavor in interspecific hybrid grape varieties: a post hoc analysis method for quantitation of volatiles and a profile of malate during berry maturation across Vitis spp. PhD Thesis, Cornell University


Progress 09/01/17 to 08/31/18

Outputs
Target Audience:Results from this project have been disseminated to a broad range of communities of interest including the general public, industry members, stakeholders, extension, and scientists through national and international conferences includingthe 69th Annual Conference of the American Society for Enology and Viticulture (ASEV) in Monterey, CA, the international Plant & Animal Genome XXVI Conference in San Diego, CA, the American Society for Enology and Viticulture-Eastern Section 43rd Annual Conference in King of Prussia, PA, the XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, and the International Congress of Plant Pathology Conference, Boston, MA. Additionally, results have been disseminated through peer-reviewed publications, trade publications, news articles, fact sheets, field days, seminar presentations, workshop presentations, stakeholder board meetings, the VitisGen2 website https://www.Vitisgen2.org/, the enology.umn.edu website, social media via Twitter, conference calls with the Advisory Panel and with co-PIs, and one-on-one communications. Specifically, targeted audiences were reached through several products and publications resulting from this project. Online webinars focused on a target audience consisting of grape scientists and grape industry members who are potential producers and users of new, disease resistant varieties and scientists and industry members interested in computer vision-based laboratory evaluation of traits. The data and research materials produced from this project targeted grape industry members and USDA funding agencies. Seminars, conferences, presentations, and workshop materials were delivered to scientists and grape industry members interested in breeding for disease resistance, management of new disease resistant varieties, and cutting-edge genotyping technologies. The databases that were disseminated were aimed at targeting project participants involved in genetic analyses or application of genetic results and all scientists who generate data on grapevines. Surveys and models that were developed targeted scientists and industry members interested in the economics of grape production. Lastly, a plant patent targeted grape industry members interested in cold-hardy, disease resistant wine grapes with excellent fruit quality. Changes/Problems:For the Breeding and Genetics Teams some small issues with leaf tissue submission and DNA extraction techniques were a problem in the early spring of 2018, but alterations in the precise protocol followed by the company hired for DNA extraction resulted in greatly improved DNA quality and all subsequent sample submissions utilized the improved protocol. For data analysis and management, a modification was made to the existing genotyping data management pipeline to keep track of the status of each sample in the genotyping process. This was necessary for a high-throughput project like this, to ensure quality and timely turnaround of results. At the FQ/Wine Center, it was planned to perform phenotyping on the same population planted in different regions. However, this was not possible because grapes from replicated field sites were not yet available. At the FQ/Table Grape Center, 2D and 3D phenotyping are now being done in collaboration with Dr. Amy Tabb, as she has extensive experience using 3D reconstruction of 2D images. What opportunities for training and professional development has the project provided?Project participants, cooperators, and visitors received on-site training in diverse topics such as: grape hybridization, propagation, and production; laboratory phenotyping of disease resistance; fruit chemistry, and physical fruit quality; genotyping and marker-assisted selection; linkage mapping and QTL analysis, genome assembly and annotation; RNASeq analysis; economic analysis and surveys; and communication and outreach. In total, these on-site training activities benefited 19 faculty, 22 technicians, 9 postdocs, 17 graduate students, 11 undergrad students, and 39 high school students. In addition, off-site training was executed via workshops, project meetings, seminars, and webinars. Undergraduate students as well as graduate students and postdocs have been trained in abiotic stress phenotyping and QTL analysis, have worked on experimental design related to understanding differences in metabolite accumulation and degradation between wild Vitis and V. vinifera, have been trained in computational methods in core genome identification, marker design, and downstream data analysis, and trained for evaluating the economic returns from new technology in agriculture. Undergraduate training focused on: mass spectrometry, automation of data collection, and evaluation of variegation in grape seedlings, learning about berry sampling and analytical approaches, and training in viticultural practices and specialized equipment. Educational efforts and outreach were conducted via webinars, the VitisGen2 website, and trade publications to educate the grape industry community on the need and potential for genetic improvement to incorporate disease resistance and quality in new varieties. How have the results been disseminated to communities of interest?Results from this project have been disseminated to a broad range of communities of interest including the general public, industry members, stakeholders, extension, and scientists through national and international conferences including, the 69th Annual Conference of the American Society for Enology and Viticulture (ASEV) in Monterey, CA, the International Plant & Animal Genome XXVI Conference in San Diego, CA, the American Society for Enology and Viticulture-Eastern Section 43rd Annual Conference in King of Prussia, PA, the XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, and the International Congress of Plant Pathology Conference, Boston, MA. Additionally, results have been disseminated through peer reviewed publications, trade publications, news articles, fact sheets, field days, seminar presentations, workshop presentations, stakeholder board meetings, the VitisGen2 website https://www.vitisgen2.org/, the https://enology.umn.edu website, social media via Twitter, and one-on-one communications. Specifically, targeted audiences were reached through several products and publications resulting from this project. Online webinars focused on a target audience consisting of grape scientists and grape industry members who are potential producers and users of new, disease resistant varieties and scientists and industry members interested in computer vision-based laboratory evaluation of traits. The data and research materials produced from this project targeted grape industry members and USDA funding agencies. Seminars, conference presentations, and workshop materials were delivered to scientists and grape industry members interested in breeding for disease resistance, management of new disease resistant varieties, and cutting-edge genotyping technologies. The databases that were disseminated were aimed at targeting project participants involved in genetic analyses or application of genetic results and all scientists who generate data on grapevines. Surveys and models that were developed targeted scientists and industry members interested in the economics of grape production. Lastly, plant patents targeted grape industry members interested in cold-hardy, disease resistant wine grapes with excellent fruit quality. What do you plan to do during the next reporting period to accomplish the goals?For the Breeding and Genetics Teams, goals for year 2 include: Further development of LIMS for more efficient streamlined automation of primer pair and sample submission and tracking information for MAS to decrease the data turnaround time. This goal will be met by simplifying submission protocols. Finish the design of the core rhAmpSeq amplicon sequencing probe set. This goal is being met by identifying core genomic regions where currently designed probes are failing to sequence. Placement of new markers in these gaps will help increase the utility of the core set. Design new core rhAmpSeq makers in genomic regions where QTL have been identified to help fine map traits. This goal will be met following the same core methods used to design the larger 2000 core probe set. The only difference will be to design probes based on the family information for the specific QTL being elucidated. Additional wild, hybrid, and cultivated genomes will be sequenced to help cover the diversity represented in the VitisGen 1 and 2 mapping families and to increase the utility of the 2000 core amplicon probe set. This goal will be met by simply continuing to sequence and analyze genomic data as it becomes available off the sequencers. Contigs of the three genotypes will be further processed to increase assembly contiguity and shared with VitisGen2 collaborators for marker discovery. The genetics team plans to initiate RNA extraction and gene expression analysis from segregating disease resistance families. These families have been identified and a major goal of year 1 was to quality check phenotypes to makes sure only the most rigorous contrasts are initiated. In year 2, we will move past QC onto bulk segregant transcriptomics. As appropriate families are determined, parental tissue will be collected for isoSeq reference sequencing as well to further increase the probability of identifying the underlying SNPs controlling contrasting phenotypes. Genes in the three genome assemblies will be annotated to serve as a reference for the full sibling transcriptomic analysis that we will conducted in the second year of the project. Future work will include collaborating with USDA-ARS scientist Edward Buckler to adopt the PHG system, and develop a new genotyping pipeline to process rhAmpSeq data. Migration of genotyping data processing pipeline to Practical Haplotype Graph (PHG) based system to improve both quality and speed of the MAS analysis pipeline. Breeding work will continue to generate and analyze combinations of fruit quality traits including plans to quantify the red-skin color of all fruiting vines in the red color population growing in Parlier, CA. Additionally, plans are scheduled to collect leaf tissues from the red color population as well as from the new PM resistance source populations for genotyping. Additional work will focus on powdery mildew resistance loci challenged with a range of strains of the pathogen. Plant samples will continue to be supplied to the phenotyping centers, and analyzed locally for additional traits of interest. Plant samples will also be supplied to the genetics team for DNA analyses leading to maps covering all 19 linkage groups. At the Missouri State Fruit Experiment Station (MSFES) efforts will continue to preserve the existing mapping populations of 'Norton' x 'Cabernet Sauvignon', 'Chambourci'n x 'Cabernet Sauvignon' and Jaeger 70/Munson x Vignoles at the Missouri State Fruit Experiment Station (MSFES) and further work will continue to construct a 'Chambourcin' linkage map with SSR markers using JoinMap 4 software. The Trait Economics Team's future plans and goals include: Administering surveys to market intermediaries and consumers to evaluate preferences with respect to varietal traits and breeding methods used on wine and table grapes. Additional plans include completing the analysis of returns to prior varietal improvements in table grapes. The Extension and Outreach Team's future plans and goals include: Continuing highlighting scientific advances from VitisGen2 through trade articles, webinars, and the VitisGen2 website. Specific topics will include economics of powdery mildew resistant varieties; challenges of durable genetic disease resistance; focus on table grape breeding and use of embryo rescue for seedless varieties; how the MN breeding program created a new cold-hardy grape industry; DNA markers for fruit quality Collaborative efforts will be made with the Breeding and Genetics Teams to contribute to the development of extension and outreach efforts. Specifically, writing content to discuss the application of technologies to a breeding program; the history of the cold-climate wine industry in the Midwest; how plant breeding for resistance does not necessarily mean "no spray" grape varieties; a focus on developing and harmonizing the traits evaluated in the grape community. Additional work will include networking with colleagues in curation of ontologies for grape genomics, and applying it to the VitisGen2 community.

Impacts
What was accomplished under these goals? Teams have worked collaboratively to address Objective 1 as follows: To date we have completed de novo assembly of 16 Vitis genomes, which has led to the construction of a Vitis pan- core-genome, identifying regions that are structurally stable and well conserved across the genus. PCR primers were developed from the core genome regions and will be used as markers in hybrid grape breeding. In collaboration with a U.S. company, IDT-DNA, we developed an improved low-cost, genotyping marker platform using rhAmpSeqTM technology, which focused on the core genome and existing molecular markers. The core genome set of 2000 amplicon sequencing markers are evenly distributed across all 19 chromosomes. Initial analyses indicate that over 90% of markers successfully returned data for marker assisted selection as well as construction of draft genetic maps. These markers have a good correlation between physical and genetic positions. To manage and analyze the new marker platform, a computational data analysis pipeline was set up to process both Ampseq and rhAmpSeq data, which includes sample and data management systems, software tools to process sequencing data, and a full pipeline from genotyping data to genetic maps or to marker-assisted selection. Across all institutional breeding programs, marker assisted selection analysis was conducted on ~6000 progeny using previously validated markers. Additionally, several wild grape genomes are in the process of being sequenced using PacBio methods. Libraries have been prepared for PacBio sequencing for V. piasezkii, V. romanetii and the PS11-5 breeding line that carries two qualitative loci (REN1 and RUN1) conferring powdery mildew resistance. For disease resistance phenotyping, we developed an automated robotic system for high-throughput phenotyping of living, unstained samples. The system is based on an XYZ robotic stage to manipulate a pre-arranged and large array of leaf disk samples. Images are captures on a high resolution 50 MP full-frame DSLR camera at 3.5X actual size. The great depth of field paired with high resolution allows a minimal number of images to be assembled to form a completely focused 3D image from which a convolutional neural network can correctly identify presence or absence of the relevant pathogen (E. necator) within 800 subsampled areas of each disk with 94.3% accuracy compared to human experts. Infection (+ or -) of each subsampled area allowed computation of infection severity on each replicate disk. The system can process up to 2000 samples per 8 hr day. The latest system will have increased sample processing speed and flexibility. Using this automated robotic system, laboratory-based powdery mildew or downy mildew phenotyping was completed for 6 VitisGen2 families and QTL analyses were completed. Tissues have been collected from four mapping families for RNAseq. These studies are designed to identify disease resistance candidate gene loci based on gene expression data. Currently, phenotypes and genotypes are being quality checked before RNA extraction is conducted. Over 700 samples from seven mapping populations and four grape breeders were received at the Cornell FQ/Wine Center (Ithaca, NY) from the 2016 and 2017 harvests. Routine protocols for analysis of acids, sugars, volatiles, tannin, and other components have been developed, and will be applied to the grape samples in the coming year. Time-resolved sampling of berries from accessions of wild V. cinerea and V. riparia species (USDA-PGRU Cold Hardy Grape Collection) and V. vinifera (Finger Lakes, NY) was performed during 2017 and 2018. Samples are in preparation for metabolic and transcriptomic analyses. At the Parlier FQ/Table Grape Center, high-throughput 2D phenotyping of cluster architecture and berry shape algorithms for analyses were developed using a subset of images from the previous year (collaborator: Dr. Amy Tabb (USDA ARS)); a high-throughput 3D reconstruction phenotyping of cluster architecture algorithm was developed (Amy Tabb); and fruit phenotyping evaluation methods for texture and juice production were developed. In addition, Breeding Team members have made extensive efforts to maintain core mapping families, and to sample and characterize them. For the latter goal of addressing the economic analysis of several key agronomic and quality traits to drive research and breeding, the Trait Economics Team has begun working on evaluating economic returns from varietal improvement in table grapes using both historical data and producer surveys. Concurrently, we are developing a set of instruments to use with consumers and market intermediaries of wine and table grapes to evaluate the attitudes towards varieties developed using conventional breeding vs. genetically edited varieties. This work has involved several conference calls with project scientists (breeders, pathologists, geneticists) as well as coordination with and guidance from the Industry Advisory Panel. Objective 2: Circa 7000 genotypes were screened for presence of multiple alleles for powdery mildew resistance, downy mildew resistance, leaf phylloxera resistance, phomopsis resistance, seedlessness, muscat character, mono- vs. diglucosides, and more. Based on the presence of favorable alleles, seedling retention during the first year of growth ranged from 29 to 44%. In some cases, both haplotypes at a locus were saved to validate markers, and the breeding and genetics teams successfully transitioned all marker screening to AmpSeq analyses. The automated imaging robot described under Objective 1 was a major technological innovation used for laboratory-based powdery mildew or downy mildew phenotyping in 6 VitisGen2 core families. This innovation enabled higher throughput than previously possible and repeated measures of live images, in contrast to previous methods that required destructive sampling for disease severity assessment. The Breeding Team also carried out phenotyping for the following traits in 2018: powdery mildew (field and lab); Phomopsis, flower sex, flowering time, time to veraison, downy mildew (including cooperative work with CSIRO and an NSF project); red skin quality on berries for future work with our red color population.; leaf trichomes; budbreak (field and greenhouse); and volatiles at anthesis, veraison and through maturity. As data are accumulated, QTL analyses will be run to locate alleles controlling trait variation. Pollen from a number of new PM resistant sources were incorporated into the breeding programs in 2018. Team members from Cornell, Minnesota, USDA-Parlier, and Davis, California, combined to make multiple crosses with five distinct combinations of the following PM resistance loci: Run1, Ren1, Ren2, Ren3, Ren4, Ren6, Ren7, Ren9, and Ren10. One cross also included three loci for downy mildew resistance (Rpv1, Rpv3, and Rpv10). These seedlings will be screened with genetic markers and selected for different combinations of resistance alleles. Additionally, crosses were performed between a new V. amurensis PM resistance source for both high quality table grapes and NDOV raisin grapes. Additional breeding work included analyses of a family segregating for all combinations of 0 to 4 resistance genes (Run1, Ren1, Ren6, Ren7), which was developed in 2016 and then genotyped and phenotyped to study the effects of gene combinations. Lines with stacked loci will be released for public use. Previous economic analyses under VitisGen1 highlighted the economic importance of powdery mildew resistance. With this knowledge, all participating grape breeders are using powdery mildew resistant germplasm and markers in their breeding programs. The impact of VitisGen2 was demonstrated to grape growers, enologists, and specialty crop researchers through a series of webinars, trade publications, the project website, workshops, field days, and peer-reviewed publications.

Publications

  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2018 Citation: Burzynski EA, Brown EJ, and Sacks GL. (2018). Sour grapes indeed! Malic acid increases in certain Vitis spp. during maturation. 69th Annual Conference of the American Society for Enology and Viticulture (ASEV), Monterey, CA.
  • Type: Journal Articles Status: Accepted Year Published: 2018 Citation: Burzynski-Chang EA, Ryona I, Reisch BI, Gonda I, Foolad M, Giovannoni JJ and Sacks GL (2018). HS-SPME-GC-MS Analyses of Volatiles in Plant Populations  Quantitating Compound � Individual Matrix Effects. Molecules. 23(10), 2436. https://www.mdpi.com/1420-3049/23/10/2436
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2018 Citation: Surya D. Sapkota, Li-Ling Chen, Shanshan Yang, Katie E. Hyma, Lance E. Cadle-Davison and Chin-Feng Hwang (2018). QTL mapping of downy mildew and botrytis bunch rot resistance in a Vitis aestivalis-derived Norton-based population. XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, July 2018. Abstract.
  • Type: Theses/Dissertations Status: Published Year Published: 2017 Citation: Adams, D. 2017. Genetic Analysis of Cold Hardiness in a Population of Norton (Vitis aestivalis) and Cabernet Sauvignon (Vitis vinifera) Hybrids. Master's Thesis, Missouri State University, Springfield, MO, USA
  • Type: Theses/Dissertations Status: Published Year Published: 2018 Citation: Sapkota, K. 2018. High Density Mapping and Quantitative Trait Loci Analysis for Fungal Diseases in Vitis aestivalis-Derived Norton. Doctoral Dissertation, Missouri State University, Springfield, MO, USA
  • Type: Theses/Dissertations Status: Published Year Published: 2018 Citation: Knuckle, C. 2018. The Identification of Intraspecific Hybrids Between Jaeger 70 x Vignoles Grapes Using SSR Markers. Master's Thesis, Missouri State University, Springfield, MO, USA
  • Type: Theses/Dissertations Status: Published Year Published: 2017 Citation: Divilov, K. 2017. Phenotypic and genetic studies of grapevine. Doctoral Dissertation, Cornell University, Ithaca, NY, USA
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Divilov, K.D., T. Wiesner-Hanks, P. Barba, L. Cadle-Davidson, and B.I. Reisch. 2017. Computer vision for high-throughput quantitative phenotyping: A case study of grapevine downy mildew sporulation and leaf trichomes. Phytopathology 107: 1549-1555. DOI: 10.1094/PHYTO-04-17-0137-R
  • Type: Journal Articles Status: Published Year Published: 2017 Citation: Fresnedo-Ram�rez, J., S. Yang, Q. Sun, L.M. Cote, P.A. Schweitzer, B.I. Reisch, C.A. Ledbetter, J.J. Luby, M.D. Clark, J.P. Londo, D.M. Gadoury, P. Kozma, and L. Cadle-Davidson. 2017. An integrative AmpSeq platform for highly multiplexed marker-assisted pyramiding of grapevine powdery mildew resistance loci. Molec. Breeding 37:145 doi: 10.1007/s11032-017-0739-0
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Divilov, K.D., P. Barba, L. Cadle-Davidson, and B.I. Reisch. 2018. Single and multiple phenotype QTL analyses of grapevine downy mildew resistance in interspecific grapevines. Theor. Appl. Genet. 131(5):1133-1143. DOI: 10.1007/s00122-018-3065-y
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Barba, P., J. Lillis, R.S. Luce, R. Travadon, M. Osier, K. Baumgartner, W.F. Wilcox, B.I. Reisch, and L. Cadle-Davidson. 2018. Two dominant loci determine resistance to phomopsis cane lesions in F1 families of hybrid grapevines. Theor. Appl. Genet. 131(5):1173-1189. DOI: 10.1007/s00122-018-3070-1
  • Type: Other Status: Published Year Published: 2017 Citation: Martinson, T. (2017). The Frozen Genetics of International Wine Cultivars. Wines and Vines, December, 2017. 28-29. Online at: https://www.winesandvines.com/columns/section/26/article/192690/The-Frozen-Genetics-of-International-Wine-Cultivars
  • Type: Other Status: Accepted Year Published: 2018 Citation: Martinson, T. (2018). Grape breeders No Longer Flying Blind. Wines and Vines, March, 2018. 64-65. Online at: https://www.winesandvines.com/features/article/196083/Grape-Breeders-No-Longer-Flying-Blind
  • Type: Other Status: Published Year Published: 2018 Citation: Martinson, T. and B. Reisch (2018). Is Europe Starting to Embrace Hybrid Wine Grapes? Wines and Vines, August 2018. 78-80. Online at: https://www.winesandvines.com/news/article/201942/Will-Europe-Embrace-Hybrid-Wine-Grapes
  • Type: Websites Status: Published Year Published: 2018 Citation: Martinson, T. and R. Kallas (2018) VitisGen2 website. http://www.vitisgen2.org/
  • Type: Book Chapters Status: Submitted Year Published: 2018 Citation: Londo, JP., Gutierrez, B., Martinez, D., Sapkota, S., Cadle-Davidson, L. From phenotyping to phenomics: Present and future approaches in grape trait analysis to inform grape gene function. The Grape Genome (Submitted)
  • Type: Book Chapters Status: Submitted Year Published: 2019 Citation: Alston, J.M., Sambucci, O. (2019) Grapes in the World Economy. Chapter for Dario Cantu and Andrew M. Walker, eds., The Grape Genome (Submitted)
  • Type: Book Chapters Status: Published Year Published: 2018 Citation: Alston, J.M., Lapsley, J.T., Sambucci, O. (2018) Grape and Wine Production in California. In Goodhue, R., Martin, P. and Wright, B., eds, California Agriculture: Dimensions and Issues). Giannini Foundation of Agricultural Economics, Berkeley, CA. Available at: https://s.giannini.ucop.edu/uploads/giannini_public/a1/1e/a11eb90f-af2a-4deb-ae58-9af60ce6aa40/grape_and_wine_production.pdf
  • Type: Journal Articles Status: Accepted Year Published: 2018 Citation: Sapkota, S.,Chen, L., Yang, S., Hyma, K., Cadle-Davidson, L., Hwang, C-F. 2018. Construction of a high-density linkage map and QTL detection of downy mildew resistance in Vitis aestivalis-derived Norton. Theoretical and Applied Genetics. doi: 10.1007/s00122-018-3203-6
  • Type: Journal Articles Status: Submitted Year Published: 2018 Citation: Fresnedo-Ram�rez, J.,Yang, S., Sun, Q., Karn, A., Reisch, B., Cadle-Davidson, L. Computational analysis of AmpSeq data for targeted, high-throughput genotyping of amplicons. Frontiers in Plant Science. (Submitted)
  • Type: Journal Articles Status: Awaiting Publication Year Published: 2018 Citation: Cadle-Davidson, L. A perspective on breeding and implementing durable powdery mildew resistance. Acta Hort (In press)
  • Type: Journal Articles Status: Published Year Published: 2018 Citation: Clark, M., Teh, S. L., Burkness, E., Moreira, L., Luby, J., Watson, Yin, L., G., Hutchison, W., (2018). QTL Identified For Foliar Phylloxera Resistance in A Hybrid Grape Population. Australian Journal of Grape and Wine Research 24(3): 292- 300.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Underhill, A., C. Hirsch, and M. Clark. 2018. Image-based phenotyping and genetic control of cluster density traits in interspecies grapevine hybrids. Poster presented at: Plant & Animal Genome XXVI, San Diego, CA. 13-17 Jan. Poster P0644.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Underhill, A., A. Diering, D. Tork, D. Freund, A. Hegeman, S. L. Teh, and M. Clark. 2018. Multiple phenotyping methods find QTL for berry color in an interspecific hybrid grape (Vitis spp.) population. Paper presented at: Taming High pH in the East. American Society for Enology and Viticulture-Eastern Section 43rd Annual Conference, King of Prussia, PA. 9-11 July
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Fennell, A., Alahakoon, D., Awale, M., Luby, J., Clark , M. Mapping the genetic architecture of grapevine bud dormancy and chilling fulfillment traits. XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, July 18, 2018. Abstract and talk.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2017 Citation: Burzynski, EA and Sacks, GL. (2017) Sour notes on the finish: malic acid behavior during ripening of wild Vitis spp. Cornell Recent Advances in Viticulture and Enology (CRAVE) Meeting, Nov 14, 2017. Ithaca, NY
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Sacks, GL. (2018) "Know your wine chem basics (and acidics) BEV-NY 2018 Conference. Mar 1, 2018. Rochester, NY.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Burzynski, E.A., B.I. Reisch, I. Gonda, M.R. Foolad, J.J. Giovannoni and G.L. Sacks. 2018. Internal standards roulette: Best practices for HS-SPME-GC-MS volatile analyses in plant populations. International Plant and Animal Genome Conference XXVI. 13-17 January 2018. San Diego, CA. Abstract
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: D. Cantu, A. Minio, Y. Zhou, A. Vondras, B. Gaut, M. Delledonne, G. Cramer, M. Massonnet, R. Figueroa-Balderas. 2018. Uncovering the wealth of grapevine genetic diversity through whole genome sequencing and assembly. XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, July 18, 2018. Invited session keynote speaker
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Kisselstein, B., Cadle-Davidson, L., Gadoury, D. Investigation of Erysiphe necator Population Structure using Amplicon Sequencing (AmpSeq) without Clonal Isolation. International Congress of Plant Pathology. July 30, 2018. Poster Presentation.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cadle-Davidson, L. Genetic tools for the study of light and circadian processes in microbial plant pathogens. International Congress of Plant Pathology. August 1, 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cadle-Davidson, L. A perspective on breeding and implementing durable powdery mildew resistance. XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, July 18, 2018. Invited session keynote speaker
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cadle-Davidson, L. Tools and strategies for breeding new grape varieties with durable powdery mildew resistance. ICVV, Longrono, Spain. April 2018. Invited Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Cadle-Davidson, L. Design and Analysis of Amplicon Sequencing (AmpSeq) for Targeted Multiplexed Genotyping. Gulf Coast Research Center, Wimauma, FL. January 31, 2018. Invited Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Reisch, B.I. 2018. Genetic resources and breeding: current status and shifting paradigms. XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, 15-20 July 2018. Invited keynote speaker.
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Hwang, C-F. QTL mapping of downy mildew and botrytis bunch rot resistance in a Vitis aestivalis-derived Norton-based population, XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, July 18, 2018. Oral Presentation
  • Type: Conference Papers and Presentations Status: Published Year Published: 2018 Citation: Fresnedo-Ram�rez, J., S. Yang, Q. Sun, J. Londo, M. Clark, B. Reisch, and L. Cadle-Davidson. 2018. AmpSeq as a tool for genetics and breeding of grapevine. XII International Conference on Grapevine Breeding and Genetics, Bordeaux, France, July 18, 2018. Abstract