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ABSTRACT YOUNG, ELISHEBA. Fruit Quality ABSTRACT YOUNG, ELISHEBA. Fruit Quality Evaluation of a Mapping Population and Single Nucleotide Polymorphic (SNP) Marker Discovery in Blueberry (Vaccinium) Species. (Under the direction of Hamid Ashrafi). Blueberry breeders at NC State University have released several elite cultivars that have contributed to the estimated ~$70 M statewide farm-gate value. Blueberries belong to the Ericaceae family and the genus Vaccinium with several subgenera or sections. Many commercially important cultivars released today including the parents of the population in the current study are derived from the species in section Cyanococcus. However, these cultivars may include introgressed genetic materials from other species of other sections that yet need to be discovered. Traditionally, selection for desirable traits is accomplished using recurrent selection through subjective field evaluations. Although a successful means of cultivar development, statistically only one in 10,000 seedlings is chosen as a cultivar which requires significant time, land, and labor resources. The task is made more difficult with increased ploidy levels. As such, there is growing interest in the development of genomic tools that blueberry breeders can use to make selections for fruit quality attributes more efficiently. Recently, a genetic linkage map has been used to identify quantitative trait loci (QTL) in a diploid population segregating for chilling requirements and cold-hardiness. However, little is known about the genetic mechanisms responsible for QTLs that control fruit quality traits like firmness, sugar content, acidity, and berry size in a tetraploid population of blueberries. As such, part of the research at the NC State blueberry breeding program involves the genotyping and phenotyping of mapping populations that segregate for fruit quality-related traits. In the current study, a tetraploid F1 population (n=344) of a cross between cv. ‘Reveille’ and cv. ‘Arlen’ (RA) was used for phenotypic evaluations. The collected phenotypic data and SNP data generated from sequence capture technology will be used for future genetic marker development, genetic map construction and QTL mapping. In the current thesis, we will only discuss the phenotypic evaluations of fruit quality-related traits in the RA population that were carried out in three harvest seasons from 2016-2018. The segregation of the traits in this population, their potential use for QTL mapping and the lessons learned will be discussed. In addition to being time and labor intensive, traditional breeding efforts to develop superior blueberry cultivars via interspecific hybridization followed by backcrossing has resulted in the development of modern cultivars that are segmental allopolyploids. The outbreeding nature of blueberry and the use of inter- and intra-specific hybridization during the past century has generated a lot of speculation about the relationship between the founder species and the modern cultivars. With the advent of next-generation sequencing (NGS) technologies, it is currently possible to uncover their interrelation at the whole genome level at a lower cost by sequencing each founder and cultivated species. In the second study, using Illumina sequencing, we re-sequenced 29 accessions at least 20X genome coverage. The 29 accessions were comprised of 18 different wild and cultivated species from 6 sections in Vaccinium that represent 18 diploids (2n = 2x = 24), 8 tetraploids (2n = 4x = 48), and 3 hexaploids (2n = 6x = 72). The 18 diploids represented 11 different species including: section Cyanococcus [V. caesariense, V. darrowii, V. elliottii, V. fuscatum, V. myrtilloides, V. pallidum, and V. tenellum]; section Batodendron [V. arboreum]; section Herpothamnus [V. crassifolium]; section Pyxothamnus [V. ovatum]; and section Polycodium [V. stamineum]. The 8 tetraploids were representative of 6 different species including: section Cyanococcus [V. angustifolium, V. corymbosum, V. formosum, and V. myrsinites]; section Hemimyrtillus [V. arctostaphylos]; and section Pyxothamnus [V. consanguineum]. The 3 hexaploids were all classified in section Cyanococcus [V. virgatum] and are known as rabbiteye blueberries. The re-sequencing data allowed for the discovery of single nucleotide polymorphic (SNP) markers within and between different groups. These SNP markers are easily adaptable to various SNP genotyping platforms that can be used in breeding programs, calculation of minor allele frequency, defining haplotype blocks and phylogenetic analysis. © Copyright 2019 Elisheba Young All Rights Reserved Fruit Quality Evaluation of a Mapping Population and Single Nucleotide Polymorphic (SNP) Marker Discovery in Blueberry (Vaccinium) Species by Elisheba Young A thesis submitted to the Graduate Faculty of North Carolina State University in partial fulfillment of the requirements for the degree of Master of Science Horticulture Raleigh, North Carolina 2019 APPROVED BY: _______________________________ _______________________________ Dr. Hamid Ashrafi Dr. Penelope Veazie-Perkins Chair of Advisory Committee ___________________________ _______________________________ Dr. Consuello Arellano Dr. Ross Whetten ___________________________ Dr. Paul Manos External Member DEDICATION To God YHWH and my family. ii BIOGRAPHY As a child, I would perform experiments with plants ultimately learning that plants were not sessile, boring creatures, bending aimlessly in the wind, obedient only to the whims of sunlight and water. Instead, these plants were meticulously crafted entities that possessed deeper self-regulation than my middle-school classes let-on. This passion for plants continued into my adulthood. I graduated in 2008 from Spelman College with a degree in Biology. While there, I took an elective in Botany course that really peaked my interest in plant sciences. I was instantly intrigued with the intricacies of plant life. This elective incorporated trips to sustainable herbal gardens during the year. In this class, I had my first exposure to the maintenance, expertise, and responsibility that horticulturalists require to have a persistent impact on their communities. I immediately began to delve deeper by researching fascinating plants in books and articles, establishing gardens on available plots of land, and signing up for nature hikes where I would identify various wild species of plants. Although I was originally a pre-medical student and even successfully completed one year of medical school out of the country, my return to the United States due financial constraints allowed me to really consider my future career path and make a decision based on passion. While ruminating on my next steps, I served in the capacity of AP Environmental Science and Biology teacher. In these roles, I found it extremely important to foster a deep understanding of sustainable crop production. As such I introduced community-garden projects to students wherever I taught. In 2014, I left teaching and decided on a career in plant sciences. In preparation, I interfaced with several botanists in Massachusetts who iii advised me about the best course of action for my career switch. One botanist in particular, Dr. Adán Colón-Carmona, was very helpful and suggested I sit in his higher-level plant physiology course at University of Massachusetts, Boston (UMASS Boston). The class and laboratory research work were so interesting that I ended up enrolling in the course. While there I was responsible for conducting an individual project that involved the expression of the Lateral Organ Boundary gene LBD10 in response to auxin in wild-type and Auxin Response Factor 7/19 double mutant Arabidopsis plants. I knew I had found a field that I loved. Upon moving to North Carolina, I asked to volunteer with Dr. Todd Wehner, a well- established cucurbit breeder at NC State University. While here, I acquired several field, greenhouse, and lab research skills that would eventually solidify my decision to become a horticulturalist. Improving the quality of the food for all communities was definitely in line with my recent career path. I applied that summer to enroll as a Master’s Student at NC State University and was accepted into the Spring 2017 cohort under Dr. Hamid Ashrafi, a highly competent blueberry breeder. Eager to begin my research, I worked as a lab technician from 2016 until the start of my Master’s program, increasing my proficiency in performing DNA extractions, library preparations, gel electrophoresis, Quanti-IT PicoGreen ds DNA assay and qPCR analysis in addition to collecting agronomic data. Under Dr. Ashrafi, I began work on two projects that attempted to map out the relationships between fruit quality traits and the QTLs that control them as well as the genetic relationship between and among several wild and cultivated species. The first project involved the measuring fruit quality related traits for QTL mapping in an F1 population of blueberry from iv a cross between ‘Reveille’ x ‘Arlen’ cultivars. Once complete, this research will bring breeders one step closer to identifying and locating QTLs associated with important traits like firmness, soluble solid content, titratable acidity, size, etc. so that selections can be made more easily for breeding. This is important because traditionally, the selection for desirable traits is accomplished using recurrent selection through subjective field evaluations. Although a successful means of cultivar development, statistically only one in 10,000 seedlings is chosen as a cultivar which
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