Understanding the genetic basis of quality-related traits contributes to the improvement of grain protein concentration (GPC), grain starch concentration (GSC), and wet gluten concentration (WGC) in wheat, a genome-wide association study (GWAS) based on a mixed linear model (MLM) was performed on the 236 wheat accessions including 160 cultivars and 76 landraces using 55K single nucleotide polymorphism (SNP) array in multiple environments. A total of twelve stable QTL/SNPs were identified to control different quality traits in this populations at least two environments under stripe rust stress; three, seven and two QTLs associated with GPC, GSC, and WGC were characterized respectively and located on chromosomes 1B, 1D, 2A, 2B, 2D, 3B, 3D, 5D, and 7D with the range of phenotypic variation explained (PVE) from 4.2 to 10.7%. Compared with the previously reported QTLs/genes, five QTLs (QGsc.sicau-1BL, QGsc.sicau-1DS, QGsc.sicau-2DL.1, QGsc.sicau-2DL.2, QWgc.sicau-5DL) were potentially novel. KASP markers for SNPs AX-108770574 and AX-108791420 on chromosome on 5D associated with wet gluten concentration were successfully developed. Phenotype of the cultivars containing the A-allele in AX-108770574 and T-allele in AX-108791420 were extremely significantly (P<0.01) higher than that of the landraces containing the G-allele or C-allele of wet gluten concentration in each of the environments. The developed and validated KASP markers could be utilized in molecular breeding aiming to improve the quality in wheat.