Date of Graduation

12-2021

Document Type

Thesis

Degree Name

Master of Science in Horticulture (MS)

Degree Level

Graduate

Department

Horticulture

Advisor/Mentor

Margaret L. Worthington

Committee Member

Renee Threlfall

Second Committee Member

Jacquelyn Lee

Third Committee Member

Leandro Mozzoni

Keywords

Blackberry, GWAS, Rubus

Abstract

The global blackberry (Rubus L. subgenus Rubus Watson) industry has experienced rapid growth during the past 15 years. Even so, many industry stakeholders report complaints from consumers and grocers stating blackberries are often too tart, too seedy, or not sweet enough for their liking. The development of molecular markers for high sweetness, low acidity, and reduced seediness would allow breeding programs to expeditiously make selection and crossing decisions in the early stages of the breeding pipeline. The objective of this study was to use a Genome-Wide Association Study (GWAS) to identify marker-trait associations, locate quantitative trait loci (QTL), and find possible candidate genes related to sweetness, acidity, and seediness in autotetraploid blackberries. A panel of 307 commercially-available cultivars and University of Arkansas System Division of Agriculture (UA) breeding selections grown at the Fruit Research Station in Clarksville, Arkansas, was phenotyped for soluble solids content (SSC) and pH in 2019, 2020, and 2021. Samples from a subset of 277 cultivars and breeding selections harvested during the summers of 2019 and 2020 were also evaluated for titratable acidity (TA), 100-seed weight, seed width to length (WL) ratio, seed area, and seeds per berry. The Rubus argutus Link. reference genome was used to design 35,054 Capture-Seq probes distributed across the genome, which were used to genotype the GWAS panel. Heritability estimates for flavor attributes concluded SSC had the lowest broad sense entry mean heritability at 61%, while pH and TA had heritabilities of 67% and 69%, respectively. Seediness attribute heritability estimates were 91% for 100-seed weight and WL ratio, 89% for seed area, and 70% for seeds per berry. Association analysis was performed in GWASpoly with 124,564 single nucleotide polymorphisms (SNPs) generated by Capture-Seq genotyping and a total of six QTL were identified. Three of the six QTL were related to flavor attributes; one for TA on Ra01, one for SSC on Ra02, and a shared QTL for TA and pH on Ra05. The remaining three QTL were related to seediness; one for WL ratio on Ra01, one for 100-seed weight on Ra03, and one for seed area on Ra05. No significant markers or QTL were identified for seeds per berry. Ten possible candidate genes for blackberry flavor attributes were identified, including an H+-ATPase 8 and a vacuolar proton-translocating pyrophosphatase associated with the QTL for TA on Ra01, a sucrose binding protein for the SSC QTL on Ra02, and two ALMT9 proteins, a MYB1, a PEPC, and three malate synthase genes for the shared TA and pH QTL on Ra05. Six seediness candidates were identified consisting of genes annotated for BRI1, CKX2, and GG3 associated with the WL ratio QTL on Ra01, an RPT2A and an AP2C1 in the QTL for 100-seed weight on Ra03, and a DA1 protein in the seed area QTL on Ra05. These results will be used to develop diagnostic markers for attributes related to sweetness, acidity, and seediness and as a training population for genomic selection.

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