In vitro studies of antioxidant potential in Rauvolfia serpentina l. under PEG induced drought
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Date
2018-04
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G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand)
Abstract
Drought affects the growth and development of plants but some medicinal plants are able to cope up with this condition by synthesizing and accumulation of antioxidants and secondary metabolite. Rauvolfia serpentina is one of them. In R.serpentina genotype Cim-Sheel and wild callus induction and proliferation was obtained in MS medium supplemented with 2, 4D (2. 0 mg l-1) and BAP (0.5 mg l-1), maximum (90 & 84 %) callus induction was recorded in minimum (22.90 and 23.00) days after inoculation with maximum dry weight (0.274 & 0.270) respectively. Highest (92.59 & 70.36 %) shoot induction was obtained in MS medium supplemented BAP (2.0 mg l-1) and NAA (0.2 mg l-1 ) in minimum (24.00 & 25.67) days after inoculation while maximum shoot length (5.17 & 4.17 cm) and shoot number (4.67 & 3.33) was also recorded in genotype Cim-sheel and wild respectively. A noval approach for in vitro flowering was initiated by using the BAP (1.0 mg l-1) and 2, 4 D (0.5 mg l-1). Among the all combination, this combination was found maximum (83.33 %) flowering induction obtained in minimum 28 days after inoculation in Cim-Sheel genotype. Drought stress (1, 2 & 3%) PEG treatment was given to the proliferated callus and regenerate shoots of both genotype Cim-sheel and wild. The effect of the drought was investigated by taking various parameters such as RWC, membrane stability, antioxidants and protein profiling. The minimum (22.37 and 21.85 %) RWC was found in proliferated callus and (21.70 and 18.84 %) was found in regenerated shoot of genotype Cim-Sheel and wild respectively in 3 % PEG after 45 days of inoculation. Electrolyte leakage and MDA content was found increase (71.23 and 80.00%) and (0.155 & 0.73 μmol/mg Fw) in 1 % PEG treatment at 45 days after inoculation. A significant increment was found in proline content under stress was recorded maximum (3.24 & 3.60 μmol/mg FW) is regenerated in leaves of genotype Cim-Sheel and wild at 3 % PEG after 45 days to inoculation as compare to control. The Phenol and the flavonoids as an antioxidant were enhanced significantly with increasing drought and days after treatment. The maximum phenol (337.56 mg/g FW & 404.22 mg/g FW) and flavonoids (41.33 & 47.41 mg/g FW) in callus and regenerated leaves were found increase under 3 % PEG at 45 days after inoculation respectively. While maximum ascorbate content (6.17 & 6.27 mg/g FW) in proliferated callus and regenerated leaves was found enhance at 3% PEG after 45 days of inoculation in Cim-Sheel genotypes of R.serpentina. The enzymatic antioxidant SOD and CAT was also increased significantly with days and time interval. Maximum SOD activity (3.27 & 3.40 unit/ mg protein) and CAT activity (22.67 and 23.23 unit/ mg protein) was recorded in proliferated callus and regenerated leaves in 3% PEG after 45 days of inoculation. For investigation of specific proteins under drought, 2DE approach was used. Total 40 proteins were detected. Out of these 19 spots were found unregulated and 11 spots down regulated and remaining had no effect of stress. Out of 19 regulated proteins two new proteins were expressed namely pyrophosphate energized vacuolar membrane pump and RNA directed DNA methylation 4 as compared to control.
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