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Subject: Botany × End date: 2017 × Start date: 2017 × Thesis Type: Ph.D ×
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    In vitro establishment of Polygonatum verticillatum (L.) All. and augmentation of its phytochemical profile using elicitors
    (G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-11) Tiwari, Tanuja; Chaturvedi, Preeti
    lants as a valuable and safe natural source of medicine have drawn much scientific and commercial attention in past few decades. Polygonatum verticillatum (L.) All., an Asparagaceae member, is an important and highly valued ‘Astavarga’ medicinal herb of North West Himalaya. Regeneration of the plant through seeds is a slow process in nature owing to epicotyl dormancy. Therefore, in the present study, an effort was made to break seed dormancy, increase seed germination potential and micropropagate P. verticillatum using epicotyl and sprouted buds. Among all storage conditions and PGRs, cold moist stratification of seeds (4±10C for 4 weeks) together with GA3 (100 mg/L) pretreatment followed by germination on GA3 (50 to 100 mg/L) supplemented MS medium was found most effective to improve seed germination. Maximum per cent germination (78±3.74) was achieved through elicitation using 50µM sodium nitroprusside (SNP). Maximum per cent bud induction (90±6.1) with maximum number of buds (7.0±0.45) and shoot length (7.10±3.18cm) was achieved from epicotyl explants inoculated in MS+TDZ(1.0mg/L)+NAA(0.5mg/L)+GA3(2.0mg/L). Microshoots inoculated in MS+KN (1.0mg/L)+IAA(0.5mg/L) and MS+IBA(1.0 mg/L) respectively showed maximum per cent microrhizome induction (95±5.0) and root induction (85±6.12). Sprouted bud explants inoculated in MS+TDZ (2.0mg/L)+NAA(1.0mg/L)+GA3(2.0mg/L) showed maximum per cent shoot induction(85±10.0). However, maximum per cent rooting (70±5.0) was achieved in MS+IBA (1.0 mg/L) and MS+NAA (1.0 mg/L)+KN(0.5mg/L). Plants raised through epicotyls and sprouted buds showed 80% and 56% survival rate respectively in garden soil. An effort was also made to investigate the effect of various chemical elicitors on phytochemical profiling of the plant. Maximum total chlorophyll content (1.67±0.05 mg g-1 fresh wt.), total phenolic content (14.71±0.47mg GAE g-1), total flavonoid content (14.66±0.17 mg QE g-1) and %DPPH radical scavenging activity (60.23±0.77) were achieved in SA (300 µM), JA (300 µL/L), SA (300 µM) and SNP (50µM) elicited plants respectively. Protein content (2.40±0.06 mg g-1 fresh wt.), peroxidase activity (1.33±0.03 unit min-1mg-1protein) and catalase activity (1.32±0.05 unit min-1mg-1protein) were augmented maximum in underground parts of JA (300 µL/L) and SNP (50 µM and 100 µM) elicited plants respectively, whereas SOD activity (4.79±0.13 unit min-1mg-1protein) was reported maximum in aerial parts of SA (300 µM) elicited plants. GC-MS analysis of differently elicited plants divulged remarkable (%) increase in medicinally important compounds. Diosgenin (anticancerous, antidiabetic), 9,12Octadecadienoic acid (z,z) (anti-inflammatory and antiarthritic), Gamma-sitosterol (antidiabetic) and Squalene (anticancerous, anti-tumor) increased to maximum viz., 68.11%, 89.67%, 78.87% and 81.82% respectively in SA treated plants. Guanosine (antiprotozoal), Palmitic acid (antioxidant), Vitamin E (antiinflammatory), 1,2-Benzenedicarboxylic acid (anti-inflammatory) increased by 53.25%, 51.68%, 83.04%, 66.50% respectively in SNP elicited plants. 2,3-dihydro-3,5-dihydroxy-6-methyl-4H-pyranone (anticancerous), Deoxy-d-mannoic lactone (antibacterial) increased by 23.56% and 66.67% respectively in JA elicited plants. The present investigation gave a complete reproducible protocol for in vitro establishment of P. verticillatum. Furthermore, elicitation study on the plant helped in augmentation and authentication of neutraceutical status of the plant.
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    Assessment of morphological diversity and biochemical parameters in some selected genotypes of Trigonella foenum-graecum L.
    (G.B. Pant University of Agriculture and Technology, Pantnagar (Uttarakhand), 2017-01) Nisha; Rao, P.B.
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    ThesisItemOpen Access
    Isolation, characterization and growth promoting attributes of plant growth promoting bacteria (PGPB) on field crops under irrigated and drought conditions
    (G.B. Pant University of Agriculture and Technology, Pantnagar (Uttarakhand), 2017-01) Dinesh Chandra; Sharma, A.K.
    Plant growth promoting bacteria (PGPB) containing 1-aminocyclopropane-1-carboxylate (ACC) deaminase has been reported to play an important role in alleviation of drought stress. In present study, eight ACC deaminase producing tolerant bacteria were isolated from a rainfed agriculture field of Kumaun region of Central Himalaya, Uttarakhand, India and evaluated for their efficiency in improving the growth of finger millet (Eleusine coracana (L.) Gaertn.), rice (Oryza sativa L.) and wheat (Triticum aestivum L.) under irrigated and drought conditions. These bacteria withstand a substrate metric potential of -1.0 MPa (30% PEG 8000) and therefore are considered to be drought-tolerant. The selected bacterial isolates were identified as Ochrobactrum sp., Pseudomonas sp., and Variovorax sp. by fatty acid methyl ester (FAME) analysis and 16S rRNA gene sequencing. The presence of acdS, the structural gene for ACC deaminase in selected bacterial strains was confirmed by the polymerase chain reaction (PCR). Glass house experiment showed that enhanced level of nutrients and antioxidant properties with the consortium of different bacterial strains. The consortium of Ochrobactrum anthropi DPC9 + Pseudomonas palleroniana DPB13 + Pseudomonas fluorescens DPB15 + Pseudomonas palleroniana DPB16 significantly increased the 1000 grain weight (9.3%, 28.9%), number of grains/panicle or spike (31.3%, 32.4%), grain yield (39.6%, 42.7%), and straw yield (21.1%, 26.9%) of rice and wheat, respectively as compared to their respective non-inoculated plants under organic field conditions. Variovorax paradoxus RAA3 treated plants maximally increase the total tillers/plant (40.0%, 31.7%), number of grains/spike (30.3%, 28.2%), spike length (26.4%, 25.8%), 1000 grain weight (17.4%, 22.0%), grain yield (28.2%, 47.4%), straw yield (11.2%, 26.0%) and harvest index (10.6%, 15.3%) under irrigated and rainfed conditions, respectively as compared to non-inoculated plant. Quantitative RT-PCR (qRT-PCR) based expression analysis of drought responsive genes (helicases and aquaporin) was also performed in response to V. paradoxus RAA3 inoculation in two varieties of wheat (HD 2967 and PBW 660) under irrigated and rainfed conditions The relative expression of aquaporin (TaTIP1;1) is significantly higher (7.26-fold) in the wheat variety PBW 660(drought tolerant), as compared to drought sensitive HD 2967 where expression level of aquaporin is down-regulated. The transcript level of WDH45 is down-regulated in both the varieties under irrigated conditions. In rainfed conditions, the relative mRNA level of wheat DNA helicase (WDH45) is significantly higher in HD 2967 (1.39-fold) than the PBW 660, whereas the expression pattern of aquaporin in PBW 660 is higher (2.70-fold) than the variety HD 2967 under rainfed conditions. This finding may imply PGPB having ACC deaminase activity could turn out to be effective inoculants for improving the growth of plants. Such promising strains could possibly be used under field conditions in rainfed environment where agriculture is exclusively dependent on rainfall.