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Abstract

Identifying the genetic basis of a trait requires complementary methods to locate and investigate how genes interact to form a phenotype. Various molecular tools are currently employed in sunflower research and breeding. I aim to expand this molecular ‘toolbox’ by adapting new methods to investigate the basis of trait inheritance. Here, I investigate the genetic basis of a hypocotyl pigmentation locus for use as a native genome engineering reporter and adapt molecular tools to construct the recombination landscape, improve genome assemblies, and genetically modify cultivated sunflower. In Chapter 2, we find that variation of hypocotyl colour of seedlings in cultivated sunflower is probably explained by a large novel insertion within the previously identified locus controlling this trait. In Chapter 3, I apply hybrid pooled-pollen sequencing using synthetic long reads for identifying recombination breakpoints with a median resolution of ~7 kbp, which has moderate correlation with an existing genetic map obtained from the same cross. This method for measuring genome-wide recombination patterns can provide an alternative approach to study how genetic and environmental factors affect the distribution of recombination events. In Chapter 4, we apply Strand-seq, a single-cell directional sequencing method, for the first time to a plant species. The resulting data was used to improve the quality of the reference sunflower assembly, providing long-range ordering and orientation and complementing other sequencing data. In Chapter 5, I aimed to develop a robust method for genetic engineering, which is currently lacking in sunflower as it is generally recalcitrant to commonly used transformation approaches. While I could not recover transgenic seeds, I identified a method suitable to transfect pollen as a vector for transformation that resulted in transient expression of a reporter gene in vitro. Together, these experiments provide new tools for research and breeding in sunflower and offer insights into the complexities of identifying gene function and characterizing trait inheritance in this crop.