A Multi-Locus Genome-Wide Association Study Uncovers Candidate Genes for Alkali Tolerance in Maize (Zea mays L.) Seedlings

Soil salt-alkalization has become a major environmental stress factor limiting the improvement of maize yield. Discovering and deploying genes related to alkali tolerance is of great significance for enhancing maize alkali resistance. Here, we employed an association panel comprising 212 maize inbred lines to identify genetic loci associated with alkali-stress tolerance at the seedling stage using the Maize6H-60K single-nucleotide polymorphism (SNP) array. Genome-wide association analysis (GWAS) using six models identified 102 significant SNPs, eight of which showed consistent colocalization across multiple models. These SNPs explained 3.35% to 20.01% of the phenotypic variation. Within the genomic regions covered by the eight co-located SNP loci, we identified a total of 56 candidate genes. Based on functional annotation and homologous gene expression analysis, eight of these genes were considered significantly correlated with alkali tolerance in maize. Subsequent qRT-PCR analysis validated that two candidate genes, Zm00001d014707 and Zm00001d041548 , significantly contribute to alkali tolerance in maize seedlings. KEGG pathway and GO enrichment analyses revealed that these genes are potentially involved in stress response and metabolic regulation pathways. The promoter regions of these genes contain regulatory elements associated with stress response and hormone signal transduction. Allelic effect analysis demonstrated that AA and CC were favorable alleles, and their pyramiding constituted a viable strategy to enhance alkali tolerance in maize. These results enhance our understanding of the genetic mechanisms of alkali tolerance in maize and establish a theoretical foundation for generating alkali-tolerant maize lines.