Grain kernel discoloration (KD) in cereal crops leads to down-grading grain quality and substantial economic losses worldwide. Breeding KD tolerant varieties requires a clear understanding of the genetic basis underlying this trait. Here, we generated a high-density single nucleotide polymorphisms (SNPs) map for a diverse barley germplasm and collected trait data from two independent field trials for five KD related traits:grain brightness (TL), redness (Ta), yellowness (Tb), black point impact (Tbpi), and total black point in percentage (Tbpt). Although grain brightness and black point is genetically correlated, the grain brightness traits (TL, Ta, and Tb) have significantly higher heritability than that of the black point traits (Tbpt and Tbpi), suggesting black point traits may be more susceptible to environmental influence. Using genome-wide association studies (GWAS), we identified a total of 37 quantitative trait loci (QTL), including two major QTL hotspots on chromosomes 4H and 7H, respectively. The two QTL hotspots are associated with all five KD traits. Further genetic linkage and gene transcription analyses identified candidate genes for the grain KD, including several genes in the flavonoid pathway and plant peroxidase. Our study provides valuable insights into the genetic basis for the grain KD in barley and would greatly facilitate future breeding programs for improving grain KD resistance.