Progress 09/01/13 to 08/31/15
Outputs
Target Audience: There are diverse target audiences for this project. The genomics portion of this research, which includes the characterization of the hull-less gene and the analysis of its effects on seed nutrient content, will be of interest to researchers studying seeds, seed coats, and secondary cell wall biosynthesis. This may include researchers studying secondary cell walls for the purpose of improving biofuels. The more applied aspect of the research, which includes breeding high-quality winter squash with hull-less seeds, will impact breeders, seed companies, food processors, and growers that work with winter squash. In particular, those companies that press squash seeds for oil, a growing market in the United States, will benefit from new squash varieties. Seed companies that market to home gardeners can also benefit, since they can promote new "dual purpose" squash varieties in which both the seeds and the high-quality flesh can be eaten. The creation and eventual release of the hull-less squash varieties has the potential to increase nutrition for the general public, as consumers of these new winter squash varieties will be able to also eat the nutrient-rich seeds. Finally, this project provided valuable plant breeding training to the Project Director (a PhD student) and the undergraduate student and other graduate student involved in the project's activities. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?This project provided training opportunities for an undergraduate student who worked on the seed nutrient analysis portion of the project as his senior honors thesis project. This student received one-on-one mentoring by the PD and her PI Dr. Michael Mazourek on experimental design, seed nutrient analysis, and statistical analysis of his results. It also provided the opportunity for him to receive advice and support regarding his goal of going to graduate school to study plant breeding. This project also provided training for one first-year graduate student, who was mentored by the PD as he learned lab and data analysis skills by working to test the candidates for the hull-less gene, which was part of the goals of this project. The PD, Lindsay Wyatt, has had extensive training and professional development as part of this project. She has received one-on-one mentoring in plant breeding methods from her PI and has gained skills in nutrient analysis from one of her graduate committee members. She has also had many opportunities for professional development, including presenting a poster and three oral presentations at conferences, planning and attending the third annual Student Organic Seed Symposium, managing work crews to achieve breeding program goals, mentoring undergraduate students, and networking with seed industry professionals. During this award period, the PD graduated with her Ph.D. in Plant Breeding & Genetics and was able to reach her goal of working as a vegetable breeder, as she was hired to work as a squash breeder at Johnny's Selected Seeds. The support of this award was critical in providing the training opportunities that prepared her for her new job, where she is now working to develop improved squash varieties that can serve to support local food systems. How have the results been disseminated to communities of interest? The results of this project have been presented to fellow plant scientists at several national and international scientific conferences (see "Products" section) and to seed companies at field days and through reports. The hull-less seed characteristic in squash as well as other interesting culinary attributes have been presented to culinary students and professors through guest lectures and field tours to expand their knowledge and interest in the diversity of vegetables available for use in preparing food. The PD's transcriptome sequencing work has been disseminated through an open-access publication and a podcast making the information from the publication more accessible to a broader audience. The PD's graduate work, supported by this award, was presented to the Cornell community through her Ph.D. defense seminar and more broadly through the publication of her dissertation. What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
1a) Use RNA-seq data to test the hypothesis that the hull-less gene is a transcription factor controlling secondary cell wall biosynthesis in the seed coat RNA-seq data derived from seed tissue from a hull-less and a wild-type squash was assembled into a transcriptome. Candidate secondary cell wall genes were identified in the data set and their expression was analyzed both throughout seed development and between the two different lines. The data clearly supported the hypothesis that the hull-less gene is a transcription factor, allowing the subsequent analysis of transcription factors in objective 1b. This also represents a major advance in knowledge about squash seed development that will be a resource for future studies of squash seeds. 1b) Evaluate hull-less gene candidates through analysis of RNA-seq data A large set of candidate transcription factors was assembled from the transcriptome data set and a much smaller group of the top candidates was selected after evaluating all candidates based on gene sequence and homology and gene expression patterns. 1c) Validate hull-less gene candidates in breeding populations An initial breeding test population was grown and seed phenotype data was collected as well as DNA samples. This will be used by a new graduate student, who is taking over the project after the graduation of the PD, to test the hypotheses generated in this study. 2a) Measure seed phytonutrient content of hull-less breeding lines The hull-less and hulled lines studied in the RNA-seq experiment were analyzed for seed size and weight, carotenoid content, tocopherol content, and protein content. We found that the hull-less seeds had more carotenoids and tocopherols than the hulled seeds, but had the same level of protein. 2b) Test the hypothesis that hull-less seeds have improved nutrient content Seed size and weight was evaluated in a F2 population segregating for the hull-less gene. We found that the hull-less seeds have a larger edible portion of the seed, making them a more efficient crop that dedicates more energy to making edible seeds. 3a) Select for improved fruit quality and shape in advanced hull-less winter squash backcross populations We conducted a trial of advanced breeding lines, evaluating their agronomics, seed yield, fruit yield, and fruit and seed quality. We identified promising lines to use for further breeding efforts and one that may be ready for release as-is. We also continued selecting in other advanced populations and have some high-quality winter squash with hull-less seeds. In addition, we made backcrosses in winter 2014 to continue improving fruit quality. This has resulted in high-quality winter squash that is close to being commercialized and making winter squash with edible hull-less seeds available to the public. We have interacted with a chef to test the quality and uses of these varieties and with a seed company to work towards their commercialization. 3b) Apply new genetic resources toward accelerating breeding progress We have not yet been able to develop a molecular marker for the hull-less gene, but the new graduate student taking over the project will continue working towards this goal. He is applying genotyping-by-sequencing with the new NextSeq platform that allows for nearly an order of magnitude more data than past technologies, and we are now able to call SNPs using the draft Cucurbita genome.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2015
Citation:
Wyatt, L., Strickler, S., Mueller, L., and M. Mazourek (2015). An acorn squash (Cucurbita pepo ssp. ovifera) fruit and seed transcriptome as a resource for the study of fruit traits in Cucurbita. Horticulture Research, 2. 1-7.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2014
Citation:
Wyatt, L., Dunn, A., Smart, C., and M. Mazourek. Breeding peppers resistant to Phytophthora capsici. Asian Solanaceous Round Table. Bangalore, India. September, 2014.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2014
Citation:
Wyatt, L., Strickler, S., Mueller, L. and M. Mazourek. Comparative analysis of gene expression networks underlying winter squash fruit quality. Cucurbitaceae. Bay Harbor, MI. October, 2014.
- Type:
Other
Status:
Published
Year Published:
2015
Citation:
Wyatt, L. "Culinary improvement in winter squash: Phenotypic and genomic approaches." Cornell Plant Breeding & Genetics Seminar Series, Ithaca, NY. April 14, 2015.
- Type:
Theses/Dissertations
Status:
Published
Year Published:
2015
Citation:
Wyatt, L. "Genomic and phenotypic analyses of vegetable disease resistance and fruit quality." Dissertation, Cornell University. May 2015.
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Progress 09/01/13 to 08/31/14
Outputs
Target Audience: The target audiences for this project are wide-ranging. The more fundamental aspects of the research, which include characterization of the hull-less gene and analysis of its effects on seed nutrient content, will be of interest to researchers studying seeds, seed coats, and secondary cell wall biosynthesis. The more applied aspect of the research, which includes breeding high-quality winter squash with hull-less seeds, will impact breeders, seed companies, food processors and growers that work with winter squash. In particular, those companies that press squash seeds for oil, a growing market in the United States, will benefit from new squash varieties. The creation and eventual release of the hull-less squash varieties has the potential to increase nutrition for the general public, as consumers of these new winter squash varieties will be able to also eat the nutrient-rich seeds. Finally, this project is providing valuable plant breeding training to the Project Director (a current PhD student) and the undergraduate student involved in the project’s activities. Changes/Problems: We have changed the focus of Objective 2 based on our preliminary data. Instead of analyzing six inbred hull-less lines, we have analyzed the two lines we are using for RNA-sequencing work and for our F2 test population. This use of one hulled and one hull-less line allows us to compare between the two. In addition, we have decided to analyze carotenoid content instead of fatty acid content because our prelimary data shows large contrasts in carotenoid content between hull-less and hulled lines, so this is a valuable analysis to perform. What opportunities for training and professional development has the project provided? This project provided training opportunities for one undergraduate student who worked on the seed nutrient analysis portion of the project as his senior honors thesis project. This student received one-on-one mentoring by the PD and her PI Dr. Michael Mazourek on experimental design, seed nutrient analysis, and statistical analysis of his results. It also provided the opportunity for him to receive advice and support regarding his goal of going to graduate school to study plant breeding. The PD, Lindsay Wyatt, has had extensive training and professional development as part of this project. She has received one-on-one mentoring in plant breeding methods from her PI and has gained skills in nutrient analysis from one of her graduate committee members. She has also had many opportunities for professional development, including presenting a poster and oral presentation at conferences, planning and attending the third annual Student Organic Seed Symposium, managing work crews to achieve breeding program goals, mentoring undergraduate students, and networking with seed industry professionals. How have the results been disseminated to communities of interest? The results of this project have been presented to fellow plant scientists at several national and international scientific conferences (see "Products" section) and to seed companies at field days and through reports. The hull-less seed characteristic in squash as well as other interesting culinary attributes have been presented to culinary students and professors through guest lectures and field tours to expand their knowledge and interest in the diversity of vegetables available for use in preparing food. What do you plan to do during the next reporting period to accomplish the goals? 1a) Use RNA-seq data to test the hypothesis that the hull-less gene is a transcription factor controlling secondary cell wall biosynthesis in the seed coat This objective has been completed and the results will be shared via a peer-reviewed publication. 1b) Evaluate hull-less gene candidates through analysis of RNA-seq data This objective has been completed and the results will be tested in objective 1c. 1c) Validate hull-less gene candidates in breeding populations An initial breeding test population was grown and seed phenotype data and DNA samples were collected in the first year of the project. This will now be used to develop molecular markers for the hull-less gene candidates and test whether any of the candidates co-segregate with the hull-less phenotype. 2a) Measure seed phytonutrient content of hull-less breeding lines This objective has been completed with some changes (see "Changes" section) and its results will be used to inform objective 2b. 2b) Test the hypothesis that hull-less seeds have improved nutrient content We found that the hulled vs. hull-less seeds differ in Vitamin E and carotenoid content, so we will test F2 seeds for these two traits. They do not differ for protein content so we do not need to test for protein in the F2. 3a) Select for improved fruit quality and shape in advanced hull-less winter squash backcross populations We are continuing our breeding efforts by increasing generations in Winter 2015, then will continue to evaluate and select in Summer 2015. 3b) Apply new genetic resources toward accelerating breeding progress Once we have identified the hull-less gene, we can develop a molecular marker for the gene and use it to select hull-less squash out of segregating populations in Summer 2015.
Impacts
What was accomplished under these goals?
Impact: Winter squash is a widely consumed vegetable crop. Although squash seeds are highly nutritious, they are usually discarded during food preparation because of their tough seed coat. One way to address this problem is by utilizing the hull-less gene, which causes squash to produce “hull-less” seeds that can be easily eaten. We have studied the hull-less gene, including how it causes the hull-less seed trait and the effect it has on seed nutrient content. Gene expression was analyzed to determine how genes turn on and off throughout seed development to create a normal seed coat, and how this pattern is different in hull-less seeds. This analysis helps us to understand how seeds develop in squash and is a step towards identifying the hull-less gene itself. We have also looked at the nutrient content of hull-less seeds as compared to normal seeds and these results will help consumers and food manufacturers to inform their choice of what type of pumpkin seeds to eat or use as an ingredient. Finally, we have made progress on developing improved hull-less seeded squash that have better-tasting fruit and also are more compact plants with higher disease resistance. This will allow farmers to produce hull-less pumpkin seeds with less waste, less pesticide use, and higher profits. Accomplishments: 1a) Use RNA-seq data to test the hypothesis that the hull-less gene is a transcription factor controlling secondary cell wall biosynthesis in the seed coat RNA-seq data derived from seed tissue from a hull-less and a wild-type squash was assembled into a transcriptome. Candidate secondary cell wall genes were identified in the data set and their expression was analyzed both throughout seed development and between the two different lines. The data clearly supported the hypothesis that the hull-less gene is a transcription factor, allowing the subsequent analysis of transcription factors in objective 1b. This also represents a major advance in knowledge about squash seed development that will be a resource for future studies of squash seeds. 1b) Evaluate hull-less gene candidates through analysis of RNA-seq data A large set of candidate transcription factors was assembled from the transcriptome data set and a much smaller group of the top candidates was selected after evaluating all candidates based on gene sequence and homology and gene expression patterns. This will allow the analysis of these candidates in breeding populations in objective 1c. 1c) Validate hull-less gene candidates in breeding populations An initial breeding test population was grown and seed phenotype data was collected as well as DNA samples. This will be used in the following reporting period when markers for the hull-less gene candidates are developed and tested in this population. 2a) Measure seed phytonutrient content of hull-less breeding lines The hull-less and hulled lines studied in the RNA-seq experiment were analyzed for seed size and weight, carotenoid content, tocopherol content, and protein content. We found that the hull-less seeds had more carotenoids and tocopherols than the hulled seeds, but had the same level of protein. 2b) Test the hypothesis that hull-less seeds have improved nutrient content Seed size and weight was evaluated in a F2 population segregating for the hull-less gene. We found that the hull-less seeds have a larger edible portion of the seed, making them a more efficient crop that dedicates more energy to making edible seeds. 3a) Select for improved fruit quality and shape in advanced hull-less winter squash backcross populations We conducted a trial of advanced breeding lines, evaluating their agronomics, seed yield, fruit yield, and fruit and seed quality. We identified promising lines to use for further breeding efforts and one that may be ready for release as-is. We also continued selecting in other advanced populations and have some high-quality winter squash with hull-less seeds. In addition, we made backcrosses in winter 2014 to continue improving fruit quality. This has resulted in high-quality winter squash that is close to being commercialized, making winter squash with edible hull-less seeds available to the public. 3b) Apply new genetic resources toward accelerating breeding progress This objective will be addressed in the second year of the project.
Publications
- Type:
Journal Articles
Status:
Submitted
Year Published:
2014
Citation:
Wyatt, L., Strickler, S., Mueller, L., and M. Mazourek. An acorn squash (Cucurbita pepo ssp. ovifera) fruit and seed transcriptome to aid the study of quality traits in winter squash. Submitted.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Wyatt, L. and M. Mazourek. Pleiotropy in Breeding for Flavor in Vegetables. American Society for Horticultural Science Annual Meeting. Orlando, FL. July, 2014.
- Type:
Conference Papers and Presentations
Status:
Published
Year Published:
2014
Citation:
Wyatt, L., Strickler, S., Mueller, L., and M. Mazourek. Analysis of the Genetic Basis of Fruit Quality in Winter Squash (Cucurbita pepo) through Comparative Transcriptome Sequencing. Plant and Animal Genome. San Diego, CA. January, 2014.
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