Rapid development and adoption of biofortified, provitamin A-dense orange maize (Zea mays L.) varieties could be facilitated by a greater understanding of the natural variation underlying kernel color, including as it relates to carotenoid biosynthesis and retention in maize grain. Greater abundance of carotenoids in maize kernels is generally accompanied by deeper orange color, useful for distinguishing provitamin A-dense varieties to consumers. While kernel color can be scored and selected with high-throughput, low-cost phenotypic methods within breeding selection programs, it remains to be well established as to what would be the logical genetic loci to target for selection for kernel color. We conducted a genome-wide association study of maize kernel color, as determined by colorimetry, in 1,651 yellow and orange inbreds from the Ames maize inbred panel. Associations were found with y1, encoding the first committed step in carotenoid biosynthesis, and with dxs2, which encodes the enzyme responsible for the first committed step in the biosynthesis of the isoprenoid precursors of carotenoids. These genes logically could contribute to overall carotenoid abundance and thus kernel color. The lcyE and zep1 genes, which can affect carotenoid composition, were also found to be associated with colorimeter values. A pathway-level analysis, focused on genes with a priori evidence of involvement in carotenoid biosynthesis and retention, revealed associations for dxs3 and dmes1, involved in isoprenoid biosynthesis; ps1 and vp5, within the core carotenoid pathway; and vp14, involved in cleavage of carotenoids. Collectively, these identified genes appear relevant to the accumulation of kernel color.

Efforts are underway for development of crops with improved levels of provitamin A carotenoids to help combat dietary vitamin A deficiency. As a global staple crop with considerable variation in kernel carotenoid composition, maize (Zea mays L.) could have a widespread impact. We performed a genome-wide association study (GWAS) of quantified seed carotenoids across a panel of maize inbreds ranging from light yellow to dark orange in grain color to identify some of the key genes controlling maize grain carotenoid composition. Significant associations at the genome-wide level were detected within the coding regions of zep1 and lut1, carotenoid biosynthetic genes not previously shown to impact grain carotenoid composition in association studies, as well as within previously associated lcyE and crtRB1 genes. We leveraged existing biochemical and genomic information to identify 58 a priori candidate genes relevant to the biosynthesis and retention of carotenoids in maize to test in a pathway-level analysis. This revealed dxs2 and lut5, genes not previously associated with kernel carotenoids. In genomic prediction models, use of markers that targeted a small set of quantitative trait loci associated with carotenoid levels in prior linkage studies were as effective as genome-wide markers for predicting carotenoid traits. Based on GWAS, pathway-level analysis, and genomic prediction studies, we outline a flexible strategy involving use of a small number of genes that can be selected for rapid conversion of elite white grain germplasm, with minimal amounts of carotenoids, to orange grain versions containing high levels of provitamin A.