Brassica napus L. is an allotetraploid plant that contains numerous boron (B) transporter genes, the functions of which are largely unknown. Here, we identified BnaC3.BOR1, a gene expressed in various tissues, including roots, stems, leaves, and floral organs. In yeast, BnaC3.BOR1 expression significantly reduces cellular B concentration. When grown in B-poor conditions, BnaC3.BOR1 mutation significantly lowered B concentrations in the xylem sap. BnaC3.BOR1 mutants exhibited stunted growth, including shorter roots, reduced shoot biomass, and leaf curling, compared to wild-type B. napus. In low-B soil, loss of BnaC3.BOR1 function resulted in shorter plant height and abnormal floral organ development with lower B levels, which ultimately reduced yield at the reproductive stage. Furthermore, BnaC3.BOR1 is expressed in multiple tissues, with specific expression in the regions below the nodes of stems. Mutants grown in B-poor soil exhibited stem cracking, and the B concentration in these cracks was lower than in wild-type plants. These findings indicate that BnaC3.BOR1 possesses the same conserved role in roots and flowers as previously discovered BOR1s, while having a distinct, specific role in stems. Our study provides a clear example of the functional diversification of B transporter homologous genes.