With the increase in global temperature and the frequent occurrence of extreme weather, the hazards of pre-harvest sprouting (PHS) tend to increase. The occurrence of pre-harvest sprouting seriously affects the yield and processing quality of wheat, thus causing huge economic losses to crop production. Identification of new PHS resistance sites and excellent genes and analysis of the mechanism for regulating wheat PHS have become an important basis for improving wheat PHS resistance. In this study, the resistance of 476 wheat materials planted in different environments was identified by whole-sprouting method in 2021, and the genome-wide association study (GWAS) was carried out based on resequencing data to find the genetic loci related to the resistance of wheat PHS. Wheat PHS in different environments showed abundant phenotypic variation with coefficients of variation of 0.64 and 0.71. The GWAS test results showed that a total of 331 significant SNP loci (-log10(P value)>5) were detected in different environments, mainly distributed in 2A, 2B, 3A, 3B, 4A, 4B, 5B, 6B, 7A, 7B, 7D, the co-detected sites are mainly distributed on 2A and 3A. The 3A locus was the most stable in the two environments. According to its physical location, it may be the R gene that controls the color of wheat grains. The significant site of 2A is also a co-localized site, combining gene annotation and transcriptomic data, we determined that this gene is a bHLH transcription factor and that the expression pattern of this gene is regulated in response to ABA. In order to explore the key genes that respond to water during wheat pre-harvest sprouting, transcriptome and metabolomics of freshly harvested grains from different resistant varieties were determined at different water absorption time. We finally screened the genes related to the pre-harvest sprouting process by WGCNA analysis. By homologous comparison, we found that the gene was related to seed dormancy in A. thaliana and affected seed germination. Therefore, we speculated that the gene might also be involved in the germination process of wheat grains.