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ThesisItem Open Access Early detection and management of white rust disease (Albugo candida) in rapeseed mustard(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-08) Gairola, Kalpana; Tewari, A.K.Among various diseases reported to occur on rapeseed mustard, white rust caused by Albugo candida is considered as one of the most important disease due to its destructive nature, wide distribution and grain yield losses of 17-34 per cent. The present investigation was carried out with the objectives of: early detection of A. candida, the cause of white rust disease; evaluation of rapeseed-mustard genotypes in field and in glasshouse (at cotyledonary and true leaf stage) and evaluation of some new fungicides for the effective management of the disease. The early detection of A. candida was done by PCR-based assay and light microscopy. In PCR based assay the primers ITS1 (3’-GAGGGACTTTTGGGTAATCA-5’) and Short ITS JV34 (3’- CGCCATTTAGAGGAAGGTGA-5’) and JV37 (3’-GTCAAGCAAAACAT-5’) were used to amplify the ITS region of A. candida and Alternaria brassicae. PCR amplification of A. candida from inoculated symptomatic and asymptomatic leaves yielded PCR products of 1200 bp and 600 bp of ITS1 and Short ITS primers, respectively whereas no bands were amplified in A.brassicae. This confirmed the presence of A. candida in asymptomatic inoculated leaves at early stage i.e. 1, 2, 3, 4, 5 and 6 DAI. In light microscopy the presence of pathogen structures were observed from inoculated symptomatic and asymptomatic inoculated leaves. This presence of pathogen structure viz. mycelium and sporangia was observed in asymptomatic leaves at early stage at 6,7,8 and 9 days after inoculation and from symptomatic leaves at 10 and 11 days after inoculation where as no fungal structure in healthy mustard leaves after staining with 1 percent cotton blue in lacto phenol and 0.4% trypan blue. A large number of rapeseed-mustard materials collected from different sources evaluated in field and in glasshouse (at cotyledonary and true leaf stage) revealed that for the confirmation of resistant sources against white rust disease it is very essential to evaluate Brassica materials first in field and then in glasshouse at both the stages i.e. at cotyledonary and true leaf stage under high disease pressure because some Brassica materials escaped from the disease in field but found susceptible in glasshouse at both the stages (EC-399299) or only at true leaf stage ( Katili local, E. sativus, Basanti and Banarasi rai, PWR-14-8, PWR-14-9, PWR-14-10, PWR-14-11, RMT-1-10-1, IC 597942 and IC265495). Among various fungicides Metalaxyl 8% + Mancozeb 64% (Ridomil MZ @ 0.25%) and a biological origin Azoxystrobin (Amistar 25 EC @ 0.1%) were found highly effective in inhibiting sporangial germination in-vitro and were found highly effective in controlling white rust disease (no occurrence of disease) in glasshouse and field in increasing grain yield and test weight followed by Propiconazole, Tebuconazole+Trifloxystrobin, Trifloxystrobin, Kresosim methyl (each at 0.1%). Garlic bulb extracts (2%) was also found effective in managing the disease even better than some old recommended fungicidesThesisItem Open Access Studies on management of Peronospora pisi Syd. the incitant of downy mildew of pea (Pisum sativum L.)(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-08) Pandey, Puja; Kushwaha, K.P.S.ThesisItem Open Access Comparative study of tomato diseases under different growing conditions(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-08) Rautela, Pankaj; Singh, R.P.ThesisItem Open Access Elucidating the role of Trichoderma in the triple combination ‘Copper-Chitosan-Trichoderma’ for the management of late blight disease of potato (Solanum tuberosum L.)(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-06) Bhardwaj, Nitish Rattan; Kumar, J.A ‘triple combination’ comprising of low dose of fungicide (CuOH), biocontrol agent (Trichoderma) and plant defence activator compound (chitosan) developed at Biocontrol laboratory, G.B.P.U.A.&T., Pantnagar and field tested over years was found very effective in the management of late blight of potato. In the present study, the ‘triple combination’ was further investigated by testing its different variants in two geographically different conditions and it was found that the ‘triple combination’ in its various forms was found very effective in managing late blight in both hills as well as plains. Its use led to reduction of disease severity, increased plant growth and increased yield that was at par with the standard recommended fungicides, viz., mancozeb, CuOH used for late blight management. Present study also revealed that spraying of the novel combination at proper concentration (CuOH 500 ppm+CS651 500 ppm+ Tri. (1%)), proper time (5 sprays at 35,45,55,65 and 75 DAS) with proper volume of water (1000 L/ha in 1st spray, 1083 L/ha in 2nd spray, 1166 L/ha in 3rd spray, 1333 L/ha in 4th spray and 1416 L/ha in 5th spray) resulted into significant gains in terms of disease severity reduction, plant growth promotion and tuber yield. Results of in-vitro studies conducted to find out interaction of CuOH-tolerant Trichoderma asperellum strains and chitosan showed that PCR amplification with the primers designed specifically to detect chitosanolytic genes viz., N-acetyl-β-D-glucosaminidase (NAG), hexosaminodase (HEXO) and chitinase (CHIT) resulted in expected bands of ~200bp, ~80bp and ~1450bp in case of NAG, HEXO and CHIT genes, respectively in both CuOH-tolerant Trichoderma asperellum strains TCMS-36 and SBIT-32, which indicated presence of chitosanolytic genes in the two strains. Expression profiling of the chisanolytic genes through real time-PCR revealed that chitosan incorporation into the media led to upregulation of different chitosanolytic genes. In TCMS-36, maximum expression of NAG (6.71±1.20) and HEXO (9.53±0.51) genes was observed at chitosan concentration of 500 ppm, while in case of SBIT32, maximum expression of NAG (2.82±1.45) and HEXO (4.84±1.09) was observed at 125 ppm chitosan concentration. However, in case of CHIT gene, both TCMS-36 and SBIT-32 showed maximum expression (3.29±0.91, 4.32±0.72 respectively) at 125 ppm chitosan concentration. Expression profiling of chitosanolytic genes at protein level through SDS-PAGE revealed the presence of typical band of approximate 93 KDa when these strains were grown in PDB media amended with chitosan (125ppm & 500ppm) and the band was absent when both Trichoderma strains were grown in absence of chitosan, indicating that the production of chitosanase enzyme was substrate specific and that the enzyme corresponded to an exo-type chitosanase (exo-β-Dglucosaminidase). Results of Gas chromatography-mass spectrometry revealed that in addition to efficient chitosanolytic enzymatic battery, these CuOH-tolerant Trichoderma strains also possessed important secondary metabolites such as 1,2-Benzenedicarboxylic acid, 2H-Pyran-2-one, Tri- methylsilyl palmitate, phenolic isomers, etc. that reportedly have antimicrobial and plant growth promotion activity. It is postulated that the CuOH-tolerant Trichoderma asperellum strains used in the triple combination interact with chitosan in a synergistic manner using its chitosanolytic machinery to breakdown chitosan into smaller oligomers that have antimicrobial, plant growth promoting and defence inducing activity. This aspect need to be further addressed in order to understand the mechanism of action of the triple combination in the management of late blight of potato.ThesisItem Open Access Variability in Gloeocercospora sorghi causing zonate leaf spot of sorghum and its management(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2017-07) Mamta; Singh, YogendraThesisItem Open Access Design, development and performance investigation of a boost converter based MPPT controller for photovoltaic systems(G.B. Pant University of Agriculture and Technology, Pantnagar (Uttarakhand), 2017-01) Jately, Vibhu; Arora, SudhaThesisItem Open Access Functional genomics for proving the existence of calcium binding proteins in finger millet and their facilitating role in calcium uptake under intestinal mimicking environment(G.B. Pant University of Agriculture and Technology, Pantnagar (Uttarakhand), 2016-08) Metwal, Mamta; Anil KumarThe science of nutritional biology has progressed in recent years to develop food-based nutraceuticals as a form of highly personalized medicine or therapeutic agent. Finger millet [Eleusine coracana (L.) Gaertn.] is a crop with potential but under-explored source of nutraceutical properties as compared to other cereals. Finger millet is a reasonably good source of calcium with up to 450 mg/100 g in the seeds, which is 5–10 times higher than other cereals. Therefore, products derived from finger millet can be utilized in bone mass development in growing children, other bone ailments in adults and for preventing osteoporosis. The nutritional significance of finger millet must be properly translated to nutraceutical development based on scientific rationales and applied to other staple crops for their possible enrichment. The present study was centered on to prove the existence of calcium binding proteins in the finger millet and their facilitating role in calcium uptake using various biochemical and molecular approaches (Transcriptomics & Proteomics), besides biomarker(s) based validation under in vitro intestinal mimicking environment. Peptide mass finger printing of Stains-All stained protein separated by native and SDS-PAGE electrophoresis identified the presence of calreticulin and calcineurin B like proteins in finger millet grains. These genes showed >90% similarity with rice genes. To further validate the presence of such proteins, transcriptome data of developing spikes was used to identify 4 calreticulin and 5 calcineurin B like protein genes selected based on high FPKM values. These were also abundantly expressed in high calcium (GPHCPB-45) compared to low calcium (GPHCPB-1) genotype as also evident from real time PCR analysis. Higher expression of such calcium binding proteins in developing spikes envisaged their role in high accumulation of calcium in plants. In order to establish the facilitating role of finger millet ingredients in calcium uptake, two preparations viz. citrate buffered extract (CBE) and in vitro gastro-intestinal digest (IGID) of finger millet flour of high grain calcium genotype were used for challenging CaCo-2 human intestinal cells. Cytotoxic analysis revealed that higher doses above 100µg/µl for CBE and 5µg/µl of IGID were toxic. The IC0, IC10 and IC75 values were 25, 75, 250µg/µl for CBE, 0.5, 1, 5µg/µl for IGID and 25, 50 and 100mM for the CaCl2 respectively. The cytotoxicity was also evaluated the presence of combination of CBE with CaCl2: 75+25, 75+50, 75µg/µl +75mM and in combination IGID with CaCl2: 25+25, 5+50, 5 µg/µl +75mM. There were remarkable changes in the morphological features of CaCo-2 cells challenged with treatments. The formation of early dome shaped structures, increase in villi formation with differential polarity indicated that finger millet ingredients induced the differentiation of human intestinal Caco-2 cells for better uptake of nutrients. The IGID showed maximum morphological differentiation even at low concentrations of 1µg/µl. The facilitation of calcium uptake was further demonstrated using fluorescence imaging techniques: UV-fluorescent microscope and fluorescence plate reader using Fura-2 AM dye. The IGID was more effective than CBE. The expression analysis of selected biomarkers (CALB, S100, ATPASE, SLC8A, TRPV) of saturable and non-saturable calcium uptake pathways using real time PCR showed differential transcript abundance and more prominent effect was observed in IGID treatment either alone or in combination with CaCl2. The results obtained under in vitro intestinal mimicking environment indicate the presence of calcium uptake facilitators in finger millet flour digested with intestinal enzymes. The most plausible explanations of such facilitating role are either the release of bioactive peptides or free accesses of calcium in IGID which facilitated the calcium uptake in human Caco-2 cells. Such molecular insight derived from nutritional biology research led to development of potential nutraceuticals which can be employed for minimizing the calcium deficiency disorders.ThesisItem Open Access Variability assessment of Albugo candida, the white rust pathogen of rapeseed mustard and evaluation of germplasm for resistance(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2016-08) Sharma, Vinod; Tewari, A.K.ThesisItem Open Access Studies on epidemiology and postharvest management of mango anthracnose(G.B. Pant University of Agriculture and Technology, Pantnagar - 263145 (Uttarakhand), 2016-08) Gupta, Supriya; Singh, K.P.