Key messageUsing QTL mapping and GWAS, two candidate genes (Zm00001d051039 and Zm00001d051147) were consistently identified across the three different environments and BLUP values. GWAS analysis identified the candidate gene, Zm00001d044845. These genes were subsequently validated to exhibit a significant association with maize gray leaf spot (GLS) resistance.AbstractGray leaf spot (GLS) is a major foliar disease of maize (Zea mays L.) that causes significant yield losses worldwide. Understanding the genetic mechanisms underlying gray leaf spot resistance is crucial for breeding high-yielding and disease-resistant varieties. In this study, eight tropical and subtropical germplasms were crossed with the temperate germplasm Ye107 to develop a nested association mapping (NAM) population comprising 1,653 F2:8 RILs, consisting of eight recombinant inbred line (RIL) subpopulations, using the single-seed descent method. The NAM population was evaluated for GLS resistance in three different environments, and genotyping by sequencing of the NAM population generated 593,719 high-quality single-nucleotide polymorphisms (SNPs). Linkage analysis and genome-wide association studies (GWASs) were conducted to identify candidate genes regulating GLS resistance in maize. Both analyses identified 25 QTLs and 149 SNPs that were significantly associated with GLS resistance. Candidate genes were screened 20 Kb upstream and downstream of the significant SNPs, and three novel candidate genes (Zm00001d051039, Zm00001d051147, and Zm00001d044845) were identified. Zm00001d051039 and Zm00001d051147 were located on chromosome 4 and co-localized in both linkage (qGLS4-1 and qGLS4-2) and GWAS analyses. SNP-138,153,206 was located 0.499 kb downstream of the candidate gene Zm00001d051039, which encodes the protein IN2-1 homolog B, a homolog of glutathione S-transferase (GST). GSTs and protein IN2-1 homolog B scavenge reactive oxygen species under various stress conditions, and GSTs are believed to protect plants from a wide range of biotic and abiotic stresses by detoxifying reactive electrophilic compounds. Zm00001d051147 encodes a probable beta-1,4-xylosyltransferase involved in the biosynthesis of xylan in the cell wall, enhancing resistance. SNP-145,813,215 was located 2.69 kb downstream of the candidate gene. SNP-5,043,412 was consistently identified in three different environments and BLUP values and was located 8.788 kb downstream of the candidate gene Zm00001d044845 on chromosome 9. Zm00001d044845 encodes the U-box domain-containing protein 4 (PUB4), which is involved in regulating plant immunity. qRT-PCR analysis showed that the relative expression levels of the three candidate genes were significantly upregulated in the leaves of the TML139 (resistant) parent, indicating that these three candidate genes could be associated with resistance to GLS. The findings of this study are significant for marker-assisted breeding aimed at enhancing resistance to GLS in maize and lay the foundation for further elucidation of the genetic mechanisms underlying resistance to gray leaf spot in maize and breeding of new disease-resistant varieties.
