Soybean (Glycine max L.) yield is critically influenced by the number of nodes on the main stem (MSN), which serves as the primary site for pods and seeds. To elucidate the genetic mechanisms underlying MSN, we conducted a multi-omics analysis integrating bulk segregant analysis sequencing (BSA-seq), phytohormone, and transcriptome profilings in a soybean mutant, LSD914, which exhibits a significantly increased MSN number compared to its wild-type parent, HN48. BSA-seq of an F2 population identified 27 candidate genomic regions spanning 2.92 Mb, primarily on chromosome 18. Within these regions, 149 genes harbored non-synonymous SNPs and 26 genes contained frameshift InDels, with functional enrichment pointing to pathways in plant hormone signal transduction and developmental regulation. Phytohormone profiling revealed a distinct shift in LSD914, characterized by down-regulation of jasmonates, salicylates, and auxins, alongside specific accumulation of cis-zeatin. Integrative transcriptome analysis identified Glyma.18G259400, a gene encoding a gibberellin-regulated protein (GmGASA32), which was consistently and significantly down-regulated in LSD914 across all developmental stages and tissues. This finding contrasts with previous reports of its overexpression promoting plant height, suggesting a nuanced, context-dependent regulatory role. Our integrated approach identifies a key set of candidate genes and highlights GmGASA32 as a pivotal node in a hormone signaling network that orchestrates soybean node number, providing valuable targets for breeding high-yield soybean varieties with optimized plant architecture.