Multi-model genome-wide association analysis reveals natural allelic variation and candidate loci for kernel phytic...

Phytic acid (PA) is the major phosphorus-storage compound in maize kernels, but its strong mineral-chelating capacity limits the bioavailability of essential nutrients in humans and monogastric animals. Although classical low-phytic-acid mutants have demonstrated the feasibility of phytate reduction, their use in breeding has been constrained by pleiotropic effects on seed viability, germination, and agronomic performance. Therefore, identifying natural allelic variation for kernel PA content in elite nutritional backgrounds such as Quality Protein Maize (QPM) is important for biofortification breeding. To our knowledge, no genome-wide association study has yet been reported for kernel PA content in maize. Here, we evaluated 249 QPM inbred lines across two environments and conducted genome-wide association analysis using sequencing-derived SNP markers. Kernel PA content showed substantial phenotypic variation, highly significant genotypic effects, and high broad-sense heritability in both environments, indicating strong genetic control. Of 11,525,180 variants initially called, 115,416 chromosomal biallelic SNPs passed initial filtering, and 26,074 high-quality SNPs were retained for population structure, linkage disequilibrium, and association analyses. Multi-model GWAS identified five high-confidence marker-trait associations on chromosomes 1, 3, 4, and 7. The strongest locus, chr3:236714544, was consistently detected across environments and analytical models and explained up to 17.7% of phenotypic variation. Allelic effect analysis confirmed significant differences among genotype classes at all five loci, with the clearest differentiation observed at chr3:236714544. LD-based haplotype analysis identified a three-SNP chromosome four haplotype block associated with kernel PA variation, with Hap2/GAC showing lower PA content. Integrated candidate-gene analysis prioritized seven putative candidate genes involved in transcriptional regulation, phosphate transport, phosphorylation-dependent signaling, seed development, and lipid/protein storage-related processes. Zm00001eb163880 emerged as the strongest candidate gene at the major chr3:236714544 locus, supported by a predicted missense variant and strong seed-related expression. Together, these results identify robust genomic loci, favorable allelic variation, and prioritized candidate genes for kernel PA content, providing a foundation for marker-assisted selection and future validation toward developing QPM maize with reduced anti-nutritional burden and improved mineral bioavailability.