Septoria tritici blotch (STB) is a destructive foliar diseases threatening wheat grain yield. Wheat breeding for STB disease resistance has been identified as the most sustainable and environment-friendly approach. In this work, a panel of 316 winter wheat breeding lines from a commercial breeding program were evaluated for STB resistance at the seedling stage under controlled conditions followed by genome-wide association study (GWAS) and genomic prediction (GP). The study revealed a significant genotypic variation for STB seedling resistance, while disease severity scores exhibited a normal frequency distribution. Moreover, we calculated a broad-sense heritability of 0.62 for the trait. Nine single- and multi-locus GWAS models identified 24 marker-trait associations grouped into 20 quantitative trait loci (QTLs) for STB seedling-stage resistance. The seven QTLs located on chromosomes 1B, 2A, 2B, 5B (two), 7A, and 7D are reported for the first time and could potentially be novel. The GP cross-validation analysis in the RR-BLUP model estimated the genomic-estimated breeding values (GEBVs) of STB resistance with a prediction accuracy of 0.49. Meanwhile, the GWAS assisted wRR-BLUP model improved the accuracy to 0.58. The identified QTLs can be used for marker-assisted backcrossing against STB in winter wheat. Moreover, the higher prediction accuracy recorded from the GWAS-assisted GP analysis implies its power to successfully select superior candidate lines based on their GEBVs for STB resistance.