ABSTRACT: Grain size is an important agronomic trait determines yield in barley, and a high-density genetic map is helpful to accurately detect quantitative trait loci (QTLs) related to grain traits. Using specific-locus amplified fragment sequencing (SLAF-seq) technology, a high-density genetic map was constructed with a population of 134 recombinant inbred lines (RILs) deriving from a cross between Golden Promise (GP) and H602, which contained 12,635 SLAFs with 26,693 SNPs, and spanned 896.74 cM with an average interval of 0.07 cM on seven chromosomes. Based on the map, a total of 16 QTLs for grain length (GL), grain width and thousand-grain weight were detected on 1H, 2H, 4H, 5H, and 6H. Among them, a major QTL locus qGL1, accounting for the max phenotypic variance of 16.7% was located on 1H, which is a new unreported QTL affecting GL. In addition, the other two QTLs, qGL5 and qTGW5, accounting for the max phenotypic variances of 20.7 and 21.1%, respectively, were identified in the same region, and sequencing results showed they are identical to HvDep1 gene. These results indicate that it is a feasible approach to construct a high-quality genetic map for QTL mapping by using SLAF markers, and the detected major QTLs qGL1, qGL5, and qTGW5 are useful for marker-assisted selection (MAS) of grain size in barley breeding.