Optimization of the Effect of Culture Medium and Sucrose Concentration and pH on the Production of Hairy Roots of Atropa belladonna L. Using the Response Surface Methodology (RSM) Under In Vitro Condition

Optimization of the culture medium of hairy roots in terms of the composition and type of ingredients in the medium is essential for proper growth and optimal biomass production. The greatest advantage of culturing Hairy roots is that they are often capable of producing secondary metabolites compared to the mother plant. In this research, transgenic hairy root induction was established through the mediation of the A7 strain of Agrobacterium rhizogenes. Molecular confirmation of hairy roots was done by PCR using rolB gene-specific primers. With response surface methodology (RSM) coding, the effect of three types of basal liquid MS media (MS, ½ MS and ¼ MS) with different sucrose concentrations (15, 30, and 45 g / L) and three pH levels (5.5, 5.8 and 6.1) on the growth of hairy root was investigated. The fresh and dry weight of hairy roots was measured after 4 weeks. The obtained results of response surface methods showed that the highest dry weight of hairy roots in the culture medium with a concentration of 0.56 of MS (≅ 1/2MS), 30 g of sucrose, and pH of 5.8. In optimum conditions, the utility value was 0.990, and the fresh and dry weight of hairy roots, was 8.58g and 0.317g, respectively. According to GC/MS results, the amount of atropine alkaloid in hairy roots grown in optimum culture medium was about 6.88%.