We isolated a novel allele associated with grain length and grain weight in wheat, TaGL3-5A-G. The TaGL3-5A-G allele frequency is low in wheat, so it has potential for breeding. Selection of large-grain wheat showing big grain sink potential and strong sink activity is becoming an important objective in breeding programs. Here, we cloned a wheat TaGL3-5A gene that was orthologous to rice GL3 and was phylogenetically clustered with both monocot PPKL1 and its expression pattern was similar to grain size change at early and middle stages of seed development. The isolated TaGL3-5A genomic sequence was 10,227 bp long and included 21 exons and 20 introns. Alignment of the TaGL3-5A sequences in Beinong 6 and Yanda 1817 showed a G/A substitution in the 11th exon (position 5946) that would lead to an amino acid change (Met/Ile). Subsequently, a KASP marker was designed based on this SNP. Genotyping of RILs showed that TaGL3-5A was located on the wheat 5AL chromosome and was colocated with a significant grain length QTL in three independent environments and mean value. Association analysis revealed that the TaGL3-5A-G allele was significantly correlated with longer grains and higher thousand-kernel weight. Haplotype association analysis indicated that TaGL3-5A-G could enhance grain traits in combination with TaGS5-3A and TaGW2-6B. The frequency of TaGL3-5A-G was higher in modern cultivars than in landraces but was still low in major Chinese wheat production areas. Additionally, the frequency of the TaGL3-5A-G allele in hexaploid wheat was slightly lower than in Triticum dicoccoides and much lower than in Triticum turgidum. Hence, T. dicoccoides and T. turgidum represent valuable resources for transferring the TaGL3-5A-G allele into common wheat, which should lead to longer grain length.