Prunella vulgaris L. hairy roots: Culture, growth, and elicitation by ethephon and salicylic acid

Hairy roots have gained popularity as an alternative source for secondary metabolites not only because of their biosynthetic and bioactive potentials, but also because of studies of experimental systems aiming to understand secondary metabolic pathways. While many secondary metabolites have been identified in Prunella vulgaris, a comprehensive understanding of the metabolic pathways has remained elusive. Here, Agrobacterium rhizogenes (ATCC15834)‐mediated hairy root induction of P. vulgaris resulted in 15–30 times more rosmarinic acid (RA) than in the intact plant. After the growth and RA biosynthesis of P. vulgaris, hairy roots were investigated over a 1‐month period, we studied elicitation by ethephon (Eth) and salicylic acid (SA) and found 200 μg L−1 Eth and 6.9 mg L−1 SA exhibited the optimum induction efficiency. These concentrations were further selected for a time‐course assay of hairy root cultures to assess exposure to these compounds over 8 days. During that period, RA reached maximum accumulations of 1.66‐fold 8 days after Eth elicitation and 1.48‐fold 2 days post‐SA addition. We also examined the transcripts of genes involved in the RA biosynthesis pathway, and found the expression of tyrosine aminotransferase (TAT) and phenylalanine ammonia‐lyase (PAL) were increased by Eth, and SA heightened the expression of TAT, 4‐hydroxyphenylpyruvate reductase (HPPR), PAL, 4‐coumaric acid CoA‐ligase 1 (4CL1), and cytochrome P450‐dependent monooxygenase (CYP). Altogether, our results indicate that the convenient hairy root culture system of P. vulgaris can provide an alternate source for RA production and would be conducive for the engineering of secondary metabolic pathways in P. vulgaris.