Genotyping-by-sequencing of Glossy Mutants

Glossy mutants are a common occurrence in Brassica oleracea L. and they have been documented in most crop varieties of the species including cabbage, kale, broccoli, and collard. Glossy phenotypes have been of particular interest to researchers due to observations that they influence insect behavior, in some cases conferring resistance to insect predation. Although glossy mutants are common, the exact mechanisms by which they occur are not documented. It is expected that such mutations might occur as a consequence of changes to a number of gene sequences associated with leaf waxiness. Glossy mutations following tissue culture generation of doubled haploid broccoli lines at the USDA-ARS-U.S. Vegetable Laboratory resulted in the development of two pairs of nearly-isogenic lines (NILs) that appear to differ only in leaf waxiness. Genetic characterization of both pairs of NILs through genotyping-by-sequencing resulted in over 600 homozygous polymorphisms between the NILs of each pair and 10 overlapping between the pairs. The closest gene to each polymorphism was identified using an annotated B. oleracea genome and then the resulting gene list was prioritized by functional relevance. The two glossy mutant lines likely arose from mutations of different genes in the wax synthesis pathway.