Kernel moisture content (KMC) at maturity is a crucial factor affecting the mechanical harvesting of maize. By utilizing multi-locus genome-wide association study (ML-GWAS), this study aims to explore significant genetic loci associated with KMC and provide insights into the genetic basis, which can contribute to the breeding and genetic improvement of maize varieties for appropriate harvesting. In this research, 205 maize inbred lines were used to measure the moisture content of kernels, husks, and cobs at maturity in four different environments. Six ML-GWAS methods, including mrMLM, FASTmrMLM, FASTmrEMMA, PLARmEB, PKWmEB, and ISIS EM-BLASSO, were applied in combination with 76,492 single nucleotide polymorphisms (SNPs) distributed throughout the genome for association analysis. Candidate genes were identified and gene annotation was performed. Phenotypic analysis showed that the coefficient of variation for KMC-related traits ranged from 10.47% to 33.90%, with a broad-sense heritability ranging from 67.39% to 81.24%. A total of 26, 15, and 23 SNP loci were significantly associated with kernel moisture content, husk moisture content, and cob moisture content, respectively. Among them, 14 significant SNP loci were identified by three or more methods, with a phenotypic contribution ranging from 1.13% to 17.21%. pLARmEB detected the most significant loci, while FASTmrEMMA detected the fewest. Based on the detection by three or more methods with an average PVE (Proportion of Variance Explained) of ≥5%, significant Single Nucleotide Polymorphism (SNP) loci Chr1_9232728, Chr4_176596174, Chr8_57716249, and Chr5_191021635 were identified. Around these loci, within a 220kb upstream and downstream range, a total of 17 candidate genes were excavated. These genes were predominantly enriched in cellular anatomical entities, metabolic processes, and cellular processes with catalytic activity. It is hypothesized that these genes may influence the moisture content of maize kernels, husks, and cobs by modulating cellular metabolism and catalytic activity.