~ 55 ~ Journal of Pharmacognosy and Phytochemistry 2017; SP1: 55-60 E-ISSN: 2278-4136 P-ISSN: 2349-8234 JPP 2017; SP1: 55-60 Shweta Singh Department of Bio-Engineering, Birla institute of Technology, Mesra, Ranchi, Jharkhand, India Priyanka Mishra College of Biotechnology, Birsa Agricultural University, Kanke, Ranchi, Jharkhand, India ZA Haider College of Biotechnology, Birsa Agricultural University, Kanke, Ranchi, Jharkhand, India Madhuparna Banerjee College of Biotechnology, Birsa Agricultural University, Kanke, Ranchi, Jharkhand, India Correspondence Madhuparna Banerjee College of Biotechnology, Birsa Agricultural University, Kanke, Ranchi, Jharkhand, India Efficacy of Rhizobium rhizogenesstrains on hairy root transformation in Stevia rebaudiana L. Bertoni Shweta Singh, Priyanka Mishra, ZA Haider and Madhuparna Banerjee Abstract Two strains of R. rhizogenes, ATCC15834 and MTCC 532 were tested for their virulence on different explants of Stevia rebaudiana. In case of R. rhizogenes strain ATCC 15834, out of different co-culture period, best results were observed after 48h treatment with maximum mean percentage of root induction on shoot tips (91.66%) as well as maximum mean number of hairy roots (20.6). In case of strain MTCC 532, 3h co-culture period was found to be best with maximum mean percentage of root induction (94.33%) and maximum mean number of hairy root (24.33). The overall study revealed that R. rhizogenes strain MTCC 532 was more effective than ATCC 15834 in terms of hairy root induction in S. rebaudiana. However, both the strains of R. rhizogenes (15834 and 532) were able to produce hairy roots in S. rebaudiana, where strain MTCC 532 was found better in terms of transformation frequency (94.33 %) as well as number of roots per explants (24.33). Keywords: Hairy roots, Rhizobium rhizogenes, Stevia rebaudiana, ATCC15834 and MTCC Introduction Many higher plants are major sources of natural products used as pharmaceuticals, agrochemicals, flavour and fragrance ingredients, food additives, and pesticides [1] . In the search for alternatives for production of desirable medicinal compounds from plants, biotechnological approaches, specifically, transgenic hairy root through Rhizobium rhizogenes infections, are found to have potential as a supplement to traditional agriculture in the industrial production of bioactive plant metabolites. These transformed roots have unique genetic and biochemical stability and faster multiplicity, which may provide continuous, reliable source of plant pharmaceuticals across all seasons. Stevia rebaudiana, widely used natural calorie-free sweetener [2, 3, 4] , belongs to family Asteraceae. The leaves of S. rebaudiana contain diterpene glycosides namely, stevioside, steviolbioside, rebaudioside A-F and ducloside A which are responsible for the typical sweet taste [5] . Stevioside is regarded as a valuable natural sweetening agent because of its relatively good taste and chemical stability. In addition diterpene, triterpenes, sterols and flavonoids are some of the non-sweet secondary metabolites that have also been identified from the leaves of Stevia rebaudiana [6, 7] . Beside sweetness, stevioside may also offer therapeutic benefits, as they have anti-microbial, anti-fungal, anti-hyperglycemic, anti-hypersensitive [8, 9] , anti- inflammatory, anti-bacterial, anti-viral, anti-yeast, anti-tumor, diuretic, and immune influencing properties [10] . The present study has been comprehended to check the efficacy of two strains of R. rhizogenes, ATCC15834 and MTCC 532 in hairy root transformation of different explants of S. rebaudiana. Materials and methods Plant material and R. rhizogenes strains: The plants were established in the medicinal garden of College of Biotechnology, Birsa agricultural University, Ranchi. The explants were taken from the aseptically grown plantlets multiplied in MS medium supplemented with BAP 2 mg L -1 , adenine sulphate 50 mg L -1 and citric acid 1 mg L -1 .The strains ATCC15834 and MTCC 532 of R. rhizogenes were obtained as a kind gift from Professor Kunal Mukhopadhyay, Dept. of Bio-Engineering, BIT Mesra, Ranchi. Bacterial culture condition and explant: R. rhizogenes strain ATCC15834 was cultured on YEB (Yeast Extract Beef extract) broth supplemented with 5 mg L -1 rifampicin and Strain MTCC 532 was grown on YEN (Yeast Extract with Sodium Chloride) broth and both were incubated in dark at 28°C for 48h. The cultures were then harvested at optical density 0.6 and pelleted by centrifugation at 5000 rpm for 10minutes at room temperature.