Shikonin and Luteolin Production in Hairy Root Culture of Perilla frutescens (L.)

Perilla frutescens (L.) Britton, a medicinal plant from the Lamiaceae family, was investigated for hairy roots induction and development of through two distinct experiments. The first experiment's initial phase examined the effects of explant type (leaf, cotyledon, hypocotyl, and stem internode) and co-cultivation duration (48 and 72 hours) on the efficiency of hairy root induction. The findings showed that the highest transformation efficiency, 66%, was attained by cotyledon explants co-cultivated for 48 hours. Agrobacterium rhizogenes strains (A4, ATCC 15834), explant age (7, 14, and 21 days old), and two medium types (MS and MS/2) were all optimized in the second phase. The results suggested that cotyledon explants with the ATCC15834 strain in MS/2 medium yielded the highest transformation efficiency, root count per explant, shortest rooting time, and maximum secondary hairy root development. In the second experiment, the impact of light on hairy root development was evaluated both on shaker incubators and FA-Bio bioreactor. Results showed that flask and light treatments produced the least biomass (4.14 ± 0.5 gr), while bioreactor and darkness treatments resulted in the highest biomass (20.96 ± 0.61 gr). PCR analysis confirmed the integration of rolA , rolB , and rolC genes in transgenic hairy roots, validating successful T-DNA integration. According to HPLC result, hairy roots had similar amounts of luteolin and higher concentrations of shikonin than perilla leaves, indicating that hairy root cultures are a good substitute for producing these important compounds. This study highlights the potential of cultured hairy roots as an effective method for producing essential secondary metabolites in Perilla frutescens .