Stem strength is an important indicator of lodging that significantly influences yield and quality in soybean. Therefore, understanding the regulatory mechanisms underlying stem strength is critical for ensuring soybean production. However, the genetic basis underlying this trait remains largely elusive. Here, we explored the key loci and regulators of stem strength by integrating quantitative trait locus (QTL) mapping with genome-wide association study (GWAS) using the recombinant inbred line (RIL) population and natural accessions. Finally, a major QTL covering an interval containing 15 genes was identified. One of these genes encodes a transcription factor related to WUSCHEL-related homeobox 4 (GmWOX4-like), a high-confidence candidate for regulating stem strength based on bulk transcriptome deep sequencing and single-cell sequencing. Anatomical analysis of residual heterozygous lines (RHLs) suggested that GmWOX4-like may influence stem strength by modulating cambium differentiation through various pathways. Natural variations in the GmWOX4-like promoter region showed significant correlation with stem strength and lodging ratio across the natural population. These findings provide valuable insights into the molecular mechanisms of stem strength and will contribute to marker-assisted selection for stem strength in soybean breeding.