Hairy root induction and polysaccharide production of medicinal plant Callerya speciosa Champ.

Abstract

A reliable and efficient hairy root induction system for the important medicinal plant Callerya speciosa was established by optimizing several parameters that affected the efficiency of Agrobacterium rhizogenes-mediated transformations. Four strains, A4, LBA9402, R1601 and C58C1, and four types of explants, hypocotyls, cotyledons, leaves and excised stems, were examined and the highest rate of transformation (68.3 %) was obtained in cotyledons using strain LBA9402. Therefore, the optimum co-cultivation time and acetosyringone (AS) concentration were evaluated for LBA9402-infected cotyledons and determined to be 2 days and 100 µM AS, respectively, resulting in 64.4 and 66.7 % hairy root transformation efficiency rates, respectively. Transgenic hairy root lines were confirmed by PCR amplification and Southern blot. The growth trend of hairy roots was “slow-quick-slow” and peaked after 30 days of culturing, with increases of 11-fold and 11.4-fold of fresh and dry weights respectively, compared with initial inoculation. Hairy roots showed the ability to synthesize polysaccharides. The effects of different media Murashige and Skoog (MS), ½ MS, WPM and B5, and carbon sources, sucrose, glucose, fructose and maltose on the growth and polysaccharide production of hairy roots were also studied. The ½ MS medium was found to be superior, resulting in the highest accumulation of biomass (10.9 g L−1 fresh weight and 1.1 g L−1 dry weight), the maximum growth index (9.96) and polysaccharide content (22.56 mg g−1 dry weight). Sucrose at 30 g L−1 was optimal for both biomass accumulation and polysaccharide production.