Establishment of Tripterygium wilfordii Hook. f. Hairy Root Culture and Optimization of Its Culture Conditions for the Production of Triptolide and Wilforine

Abstract

In order to solve the shortage of natural Tripterygium wilfordii Hook. f. plant resource for the production of the important secondary metabolites triptolide and wilforine, hairy roots were induced from its root calli by Agrobacterium rhizogenes. Induced hairy roots not only could be maintained and grown well in hormone-free half-strength Murashige and Skoog medium but also could produce sufficient amounts of both triptolide and wilforine. Although hairy roots produced approximately 15% less triptolide than adventitious roots and 10% less wilforine than naturally grown roots, they could grow fast and could be a suitable system for producing both secondary metabolites compared with other tissues. Addition of 50 micrometer methyl jasmonate (MeJA) could slightly affect hairy root growth, but dramatically stimulated the production of both triptolide and wilforine, whereas 50 micrometer salicylic acid had no apparent effect on hairy root growth with slightly stimulatory effects on the production of both secondary metabolites. Addition of precursor nicotinic acid, isoleucine, or aspartic acid at the concentration of 500 micrometer had varying effects on hairy root growth, but none of them had stimulatory effects on triptolide production, and only the former two had slightly beneficial effects on wilforine production. The majority of triptolide produced was secreted into the medium, whereas most of the produced wilforine was retained inside of hairy roots. Our studies provide a promising way to produce triptolide and wilforine in T. wilfordii hairy root cultures combined with MeJA treatment.

Cloning, Expression Analysis and Functional Characterization of Squalene Synthase (SQS) from Tripterygium wilfordii

“…The different fresh tissues of T. wilfordii were collected from the plants grown in the greenhouse of Northwest A & F University, China. The hairy root cultures were established as described previously [ 41 ]. The hairy root of T. wilfordii were cultured in 250 mL Erlenmeyer flasks containing 100 mL of Murashige-Skoog liquid medium, 30 g/L sucrose.…”

Elicitation: A biotechnological tool for enhanced production of secondary metabolites in hairy root cultures

“…JA, MJ, SA, and ASA have been recognized as another class of signal transducer in elicitation of plant defense responses [30,42,[65][66][67][68][69][70][71][72][73][74]. Application of MJ, JA, SA, ASA, and other signal molecules may or may not inhibit growth and thus effect biomass accumulation during the enhancement of SMs production or vice-versa [8,42].…”

Biosynthesis of Diterpenoids in Tripterygium Adventitious Root Cultures

“…Thus, our tissue cultures are an excellent experimental system for further pathway elucidation. Cultures of Tripterygium hairy roots, adventitious roots, or cell suspensions all have the advantage of accumulating abietane diterpenoids at high concentrations (Miao et al, 2013;Zhu et al, 2014) and, in comparison with the unsustainable harvest of roots (where the concentrations of these specialized metabolites are very low), should be considered as commercial sources. We demonstrate here that diterpenoids are secreted into the culture medium of T. regelii adventitious root cultures, which significantly reduces the complexity of the matrix for extraction.…”

“…Similarly, hairy root cultures were initially employed for phytochemical investigations (Nakano et al, 1998). It was recognized only recently that triptolide concentrations produced by tissue cultures (up to 0.15% of dry weight; Miao et al, 2013Miao et al, , 2014Zhu et al, 2014) far exceed those reported for roots.…”