Study of factors affecting hairy roots induction and Rosmarinic Acid production in Dracocephalum kotschyi Boiss

Abstract: The flavonoid compounds in the Dracocephalum kotschyi has anti-cancer properties and using various methods such as hairy roots production can used to produce valuable secondary metabolites. In this study, the effect of explant type (cotyledon, hypocotyl, leaf and internode) and explants age (1 week leaf, 2 weeks leaf and 4 weeks old leaf) on hairy roots induction percent were surveyed. The transgenic status of hairy roots was confirmed by PCR using rolB genes-specific primers. In order to find optimum culture media for hairy roots growth and proliferation, four types of liquid basal media including, MS, 1/2MS, 1/4 MS and B5 and UV-B elicitor effects in three levels (3 min, 6 min and 9 min) on biochemical characteristics and Rosmarinic acid (RA) contents were investigated. The maximum hairy roots induction was obtained in 1 week leaves old (56.66 %); cotyledon explants (36.66 %), respectively. The highest fresh (4.34 gr) and dry weight (0.31 gr) and lowest fresh (1.04 gr) and dry weight (0.10 gr) was recorded in 1/4 MS and B5 media, respectively. The maximum and minimum antioxidant activities (92.33% and 63 %) were obtained in UV-B exposure time to 6 min radiation and non-transformed hairy roots. High performance liquid chromatography analysis revealed that the production of RA in UV-B (6 min) elicited hairy roots of D. kotschyi was almost 2.5-fold higher than that of the non-transformed roots (34.9 µg /mg DW compared to 13.8 µg /mg DW).