Induction of hairy roots by Agrobacterium rhizogenes in relation to L-theanine production in Camellia sinensis

Abstract

SummaryTheanine is a functional compound synthesised in roots of Camellia spp., especially tea (C. sinensis). Hairy roots were induced by two strains of Agrobacterium rhizogenes (15834 and LBA9402). The amino acid, catechin and caffeine compositions of hairy roots were determined by HPLC and compared with roots grown from tea seed. The results showed that the most suitable conditions to induce hairy roots were to use stem segments cut from 70 d-old plantlets, co-cultivated on YMB (yeast, mannitol broth) medium for 2 d, then induced on MS (Murashige and Skoog) medium for 4 weeks. Induced roots grown on LG0 medium had 23.12 mg theanine g–1 fresh weight, which was 23% higher than in roots grown from tea seed. Low concentrations of catechin, caffeine and epicatechin were detected in both root samples. Polymerase chain reaction (PCR) confirmed that the genes rol A, rol B and rol C of A. rhizogenes had been transferred into the DNA of the hairy root cells. The integrated genes for plant hormone synthesis from A. rhizogenes could increase hormone levels in the induced hairy roots, which in turn stimulated root growth and theanine synthesis. Hairy roots with higher concentrations of theanine, and lower levels of catechins and caffeine, are considered to be a good source for extracting L-theanine which is used as an active ingredient in food and medicine.