Targeted metabolic engineering of key biosynthetic genes improves triptolide production in Tripterygium wilfordii hairy roots.

The overexpression of key biosynthetic genes involved in triptolide production through a metabolic engineering strategy significantly enhanced triptolide accumulation in Tripterygium wilfordii hairy roots. Triptolide, the representative bioactive compound in Tripterygium wilfordii, is renowned for its potent insecticidal and pharmacological properties. In order to increase the production of triptolide, this study overexpressed several key enzyme genes related to its biosynthesis in T. wilfordii hairy roots. Specifically, the content of triptolide in hairy roots overexpressing TwTPS9 and TwTPS27 individually was found to be 1.60-fold and 1.42-fold that of the control, respectively. Co-expression of both TwTPS9 and TwTPS27 resulted in significant increase in triptolide levels, reaching approximately 2.72 times that of the control. Furthermore, overexpressing TwGGPPS and TwDXS on the basis of the double gene overexpression led to the highest triptolide production, with a yield of 12.83 mg/L, increasing to 3.18-fold compared to the control. This study offers valuable examples into the efficient biosynthesis of triptolide and is expected to lay a foundation for future industrial-scale production by mitigating its resource constraints through metabolic engineering.