Pre-harvest sprouting (PHS) causes significant losses in wheat yield and quality worldwide. Previously, we cloned a PHS resistance gene, TaPHS1, and identified two causal mutations for reduced seed dormancy (SD) and increased PHS susceptibility. Here we identified a novel allelic variation of C to T transition in 3'-UTR of TaPHS1, which associated with reduced SD and PHS resistance. The T allele occurred in wild wheat progenitors and was likely the earliest functional mutation in TaPHS1 for PHS susceptibility. Allele frequency analysis revealed low frequency of the T allele in wild diploid and tetraploid wheat progenitors, but very high frequency in modern wheat cultivars and breeding lines, indicating that artificial selection quickly enriched the T allele during modern breeding. The T allele was significantly associated with short SD in both T. aestivum and T. durum, the two most cultivated species of wheat. This variation together with previously reported functional sequence variations co-regulated TaPHS1 expression levels and PHS resistance in different germplasms. Haplotype analysis of the four functional variations identified the best PHS resistance haplotype of TaPHS1. The resistance haplotype can be used in marker-assisted selection to transfer TaPHS1 to new wheat cultivars.