Abstract: Galactinol synthase（GolS） is a key enzyme in the biosynthesis pathway of raffinose family oligosaccharides（RFOs）. In order to explore the regulatory mechanism of soybean GmGolS under heat stress, we used real-time fluorescence quantitative PCR to detect GmGolS expression under heat stress, amplified GmGolS promoter sequence by PCR from soybean genomic DNA, constructed GmGolS promoter into plant expression vector pCAMBIA1301 and transformed into tobacco. GUS histochemical staining and real-time fluorescence quantitative PCR were used to detect the heat-induced activation activity of GmGolS promoter. The results showed that the expression of GmGolS was significantly induced by heat stress. The 1 739 bp GmGolS promoter sequence contained one auxin response element, one drought-induced MYB transcription factor binding site, two anaerobic-induced elements, one defense and stress response element, one abscisic acid response element, one jasmonic acid response element and one anoxic-induced element. We obtained three GmGolS promoter transgenic tobacco plants successfully. The activation activity of GmGolS promoter could be induced by heat stress.