Induction of hairy roots and plant regeneration from the medicinal plant Pogostemon Cablin

Abstract

An efficient transformation system for the medicinal and aromatic plant, Pogostemon cablin Benth was developed by using agropine-type Agrobacterium rhizogenes ATCC15834. Hairy roots formed directly from the cut edges of leaf explants or via callus stage 8 days after inoculation with the bacterium. The highest frequency of leaf explant transformation by Agrobacterium rhizogenes ATCC15834 was about 80% after infection for 25 days. Hairy roots grew rapidly on plant growth regulators (PGRs)-free Murashige and Skoog (MS) or 6,7-V medium and had characteristics of transformed roots such as fast growth and high lateral branching. The PCR amplification showed that rol genes of Ri plasmid of A. rhizogenes were integrated and expressed into the genome of transformed hairy roots. The hairy root line, PL6, grew very slowly in the first 8 days, then grew very quickly between day 8 and day 24. The optimum medium for callus induction of hairy roots consisted of 2.0 mg l−1 benzyladenine (BA) and 0.1 mg/l α-naphthaleneacetic acid (NAA); while optimum medium for adventitious shoot regeneration from these cultures consisted of 0.1 mg l−1 BA and 0.1 mg l−1 NAA. Adventitious shoots could be rooted on 1/2MS. Southern blot analysis confirmed that rol genes of TL-DNA of Ri plasmid was integrated with at least three copies into the genome of hairy roots- regenerated P. cablin plants. The results presented provide a solid foundation for production of patchouli essential oil from hairy roots or its regenerated plants and also provide possibilities for utilization of artifical polyploidization or chemical mutation of hairy roots for improving germplasm and breeding of a new cultivar of P. cablin.