Plant secondary metabolites are a large variety of organic compounds which are considered as the end products of plant metablism. These compounds do not play vital roles in producing plants and their production is a mechanism of compatibility.
However, human is interested in these compounds, because it uses them in many aspects of its industrial life. Plant secondary metabolites are used as perfumes, find chemicals, natural colors, food additives, natural pesticdes and therapeutics. Plants producing medicinal compounds are called medicinal plants. Among medicinal plant species, genera of Scopolia, Datura, Duboisia, Atropa and Hyoscyamus from the family of Solanaceae, produce tropane alkaloids which are valuable therapeutics. Because of high sensitivity of solanaceous plants to pests and diseases and manifold problems of growing them in vivo, many attempts have been focused on developing a powerful alternative method for producing tropane alkaloids in vitro. Callus, cell suspension and normal root cultures of producing plants have been studied extensively in many experiments. There are some major problems by using these cultures to produce tropane alkaloids in vitro, including instability in growth and biosynthesis and low yield. Thransformed hairy roots induced by Agrobacterium rhizogenes in these plants, have considerable adventitious such as rapid growth in hormone free media which makes them valuable and promising materials to produce alkaloids in vitro. Among tropane alkaloid producing plants, most attention have been foused on hairy root cultures of Hyoscyamus muticus (Egyption henbane) and Datura stramonium.
In this research, with the aim of studying growth rate and stability of transferred roots and their biosynthetic potential, leaf and nodal segments of H. muticus were inoculated by A. rhizogenes strains LBA 9402 and A4. Where hairy root were generated, They were cut and transferred to liquid Bo or MS media. Normal root cultures also were established at same time. To confirm transformation, PCR reactions for amplification of specific fragents of rolB and rolC genes of A. rhizogenes carried out on DNA of hairy root. Hairy and normal root cultures were subculture for 7 months. For growth rate analysis, dry weight of harvested root from each culture after any subculture, were measured. Hyoscyamin and scopolamine content of each dry root clamp were measured separately by HPLC. Finally, Data were analyzed by statistical methods.
A considerable biosynthctic and growth stability of hairy root in hormone free cultures was observed, whereas, normal root failed to grow in these media after two months. There was significant variation in alkaloid content and growth rates of different hairy root clones. However, growth rate of hairy roots was four times higher than normal roots, where, there was no significant difference between the alkaloid content of normal and hairy roots. Increase in biomass production without decreasing in alkaloid content, along with considerable growth stability in hairy root cultures of H. muticus, encourages studies on these hairy roots to develop an effective system for in vitro production of tropane alkaloids.