Drought adversely affects crop growth, development and yield. This study assessed genetic variation in a panel of wheat lines to water deficit and identified the QTLs governing this relationship via GWAS. A panel of 290 lines of the Wheat Association Mapping Initiative (WAMI) population were evaluated under well-irrigated and drought-stressed conditions. Stress tolerance indices were calculated to assess the response of wheat genotypes to water deficit. GWAS was performed using 15,737 SNP markers and six phenotypic traits. Evaluated traits were significantly affected across environments indicating variations in response to drought and ambient conditions. Heritability degrees were moderate to high and ranged from 0.39 to 0.93%. GWAS identified 205 significant marker-trait associations for the six studied phenotypic traits under well-irrigated and drought stress conditions. A positive correlation was found between drought stress index (STI) and plant grain yield (PGY) under both well-irrigated and stressed conditions. A linear relationship between STI and PGY under stressed conditions (PGYs) was observed. STI is the best index for prediction of high yielding genotypes. The WAMI population is a valuable source for improving drought tolerance in wheat. The study revealed significant marker–trait associations with a high degree of genetic diversity underlying different mechanisms for adaptation to drought-prone environments.