High-throughput RNA-seq technology was used to explore the gene regulation mechanism under high temperature and low dissolved oxygen stress. Overall design: In the control group, the temperature was maintained at 18°C and the dissolved oxygen was maintained at 3mg/L. In the high temperature group, heating rods were used to raise the temperature to 26°C. In the low oxygen group, nitrogen was injected into the water to make the dissolved oxygen reach 3mg/L. In the combined stress group, the temperature was 26°C and the dissolved oxygen was 3mg/L. After 6h of treatment, the intestines of Apostichopus japonicus were removed, frozen in liquid nitrogen, and then placed at -80°C until the end of analysis.