Establishment of Persicaria minor hairy roots and analysis of secreted β-caryophyllene in medium broth

Abstract

A method for hairy root induction of Persicaria minor, was developed and investigated, and a qualitative evaluation of the sesquiterpenes in the culture medium was performed. The transgenic status was confirmed by PCR analysis using rolB and virD specific primers. The efficiency with which four different strains of Agrobacterium rhizogenes (A4, ATCC43056, ATCC15834 and ATCC13333) and different concentrations of acetosyringone induced hairy roots from in vitro leaf segments of P. minor was evaluated. Strains A4 and ATCC13333 were able to induce hairy root in an average of 6 days of co-cultivation and exhibited significantly higher transformation efficiencies, 20 and 21.3 %, respectively, compared to the other strains in the presence of 100 µM acetosyringone. Half-strength Murashige–Skoog basal medium supplemented with Gamborg vitamins supported the greatest increase (35-fold) of growth index of hairy roots relative to the initial weight after 8 weeks of culture. GC–MS results showed that β-caryophyllene was the main sesquiterpenes detected and was quantified with standards, with total peak areas of 7.934. Results suggested that hairy roots secreted sesquiterpenes into the medium while preserving viable hairy roots as a source of specialized metabolites.