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Birsa Agricultural University, Ranchi
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ThesisItem Open Access Protocol Development of In- Vitro Cultivation of Bamboo ( Dendrocolamus As per )(Birsa Agricultural University, Ranchi, 2002) Peddy Srikanth; Z.A. HaitherBamboos are perennial, woody and evergreen monocotyledonous arborescent grasses belonging to the family Poaceae (Graminae) and Sub-family Bambusoideae. There are about 75 genera and 1250 species of bamboos. Dendrocalamus asper is one of the economically important and widely cultivated bamboo species. The tender shoots of this plant can be consumed as food and is a good source of foreign exchange to various countries. Mature culms of this plant are used for pulp and paper manufacture. Bamboos are propagated artificially by different methods, including through seed. But utilization of seeds as propagating material is difficult and unreliable due to long and unpredictable flowering habit, short dormancy period of seed, poor viability, inborn microbial infestation, poor seed set during off-season flowering, seed sterility and large scale. Consumption of seeds by rodents and wild animals. The vegetative methods, on the other hand, are costly, lobour intensive, cumbersome and time. Consuming. These vegetative propagates are bulky, difficult to transport to distant places and their survival rates are also not very high. This limits large scale cultivation of bamboos in general. Under the situation, propagation through tissue culture seem to be a viable method for large scale propagation of the bamboo species. Therefore the present project was undertaken to establish a protocol for in-vitro propagation of Dendrocalamus asper. In the present study nodal segments (3-4cm) with axillary buds from young juvenile mother plant was used as explants. Surface sterilization using 0.1% (w/v) mercuric chloride (Hgcl₂) for 10 minutes followed by 3-4 times subsequent washing with sterile distilled water proved the best as it resulted the highest percentage (92.68%) of bud break after two weeks. The sterilized nodal segments were cultured aseptically on MS medium supplemented with 0-15 mgl-1 BAP and maximum shoot proliferation. (14-15 shoots per propgule) was achieved on medium supplemented with 12mgl¹ BAP. These proliferated axillary shoots were excised and subcultured on MS liquid medium +3 mgl BAP for the first two subcultures to increase the number of shoots. The shoot multiplication was achieved on both MS solid as well as liquid medium supplemented with 1-5 mgl¹! BAP Highest rate of shoot multiplication (fold) i.e., 15.77 was obtained on MS liquid medium supplemented with 3 mg l-¹ BAP in four weeks. MS solid medium supplemented with 3 mgl¹ BAP resulted only 8.55 fold. Incorporation of NAA (0.2-1.0 mgl ¹) to the medium along with BAP did not increase the rate of shoot multiplication and shoot length but it resulted in better quality erect shoots. MS medium in its full strength (1x) was found to be the most effective basal nutrient medium for shoot multiplication. The studies on sucrose concentration in the medium showed that 3% sucrose was essential for rapid multiplication of shoots. The effect of pH reflected that shoot multiplication occured even on acidic medium and highest rate of shoot multiplication (15.88) was obtained at pH 5.8. A regular subculture cycle at an interval of 4 weeks resulted in healthy cultures devoid of brown leaves and high rate of shoot multiplication. For in-vitro root regeneration on MS medium supplemented with 10mgl-¹ IBA yielded 90% rooting, 19.66 roots per propagule in four weeks, while 3 mg NAA supplemented MS medium resulted 91.66% rooting with 10 roots per propagule NAA resulted short roots while IBA resulted long roots. Addition of BAP (0.1-0.5 mgl ¹) to the rooting medium, neither enhanced root regeneration percentage nor improved the number of the in-vitro roots The cultured plantlets were successfully hardened under high humidity on sterilized soil sand FYM(1:1:1) mixtured with 1/2 strength MS nutrient medium irrigations (without organics).ThesisItem Open Access Protocol Development of invitro Clonal Propagation of Orchid ( Vanda Spa)(Birsa Agricultural University, Ranchi, 2002) Ranjeet Kumar Sinha; Z.A HaitherOrchids, one of the most beautiful group of flowering plants belong to the family Orchidaceae (Monocotyledons). The exquisite beauty of Orchid flowers due to brilliance in colour, remarkable range of sizes, manifold shapes, and variation in the form and wide range of distribution has aroused highest admiration throughout the world. The Orchid comprises about 800 genera with around 35,000 species. In India, about 1300 species of Orchids are found in Himalayas and others scattered in eastern and western Ghats. A vast majority of Indian Orchids are confined to mountain where they are distributed from base of hill to the elevation of 4300 m above mean sea level in climates ranging from tropical to temperate. Orchids are terrestrial, epiphytic, lithophytic or saprophytic but no Orchid is aquatic. The cut flower trade of Orchid involves 3% internationally. Major suppliers, like, Thailand, Netherlands and Singapore export flowers worth of US $ 80.0, 77.4 and 20.0 millions, in order per year. Due to their alkaloid, flavanoid, glycosides and other phytochemical constituents Orchids have high therapeutic value. The flower juice of Vanda coerulea is used to cure eye diseases. Cymbidium elegans, Cymbidium pubescens, Epicactic latifolia are used as local medicines. for treatment of nervous disorders. Orchids are also used in many countries as food or for making refreshing drinks. Unfortunately the natural population of Orchid is fast declining due to excessive collection and over harvesting by traders and botanical explorers. So there is need to cultivate and conserve the endangered Orchids. The conventional method of propagation is tedious and time taking. The alternative means of propagation is in vitro clonal propagation. Keeping this in mind the present experiment. on developing a viable protocol for in-vitro clonal propagation of Orchid (Vanda Miss Joaquim) was undertaken. The explant, like, shoot apex and shoot node were washed with detergent and teepol and then sterilized with 0.2% mercurio chloride for 10 minutes. The explants were cut in small pieces under laminar flow hood and subsequently inoculated in Murashige and Skoog (1962) medium modified with different plant growth regulators. The inoculated materials were cultured under aseptic condition at 25+2°C with 16 hours photoperiod of 3000 lux. The medium containing 2% sucrose, 2 mgl¹ BA+ 0.2 mgl¹ NAA was best for shoot node culture and developed 10 shoots/node and 4 leaves per shoot. Protocorm like bodies were developed in cytokinins combination. The combination 1 mgl BA + 0.3 mgl kinetin proved better for getting higher number of buds. However, 7.67 buds/node were found with 1mgl kinetin + 0.1 mgl¹ 2,4-D in around 46 days. It is worthy to note here that 2 mg1¹ kinetin in absence of 2,4-D yielded 7.56 buds/node which is statistically at par with the combination treatment 1mgl kinetin + 0.1 mgl 2,4-D. The shoot apex culture gave significant results on MS. medium supplemented with 2 mgl¹ BA + 0.5 mgl'¹ NAA, 1 mgl¹ BA + 0.2mgl kinetin and 1mgl kinetin + 0.1 mgl 2,4-D. Sub-culturing of plantlet on 2 mgl¹ BA and 0.5 mgl¹ NAA gave about 70-100 shoots. Best result on rooting was achieved on MS medium supplemented with 1 mgr¹ IBA+ 0.5 mgl¹ NAA, 1 mgr¹ NAA + 0.1mgl¹ BA. The maximum root length (49.5 mm and 60 mm) was obtained on medium supplemented with 1 mgr¹ NAA+ 0.1mgr¹ BA and 2 mgl¹¹ NAA + 1 mg/¹¹ IBA respectively. The cultured shoots were hardened successfully in pots containing bark, brick pieces and charcoal in 1:1:1 ratio.ThesisItem Open Access SoybPeroatne [iGn lcyhcainrea cmtearxi z(aLt.i)o Mn efrrorimll] R frhoizmo baicuimdic is soolailste osf oSft ate of Jharkhand by utilizing immunological approach(Birsa Agricultural University, Ranchi, 2023) Srishti Kandulna; Himanshu Dubey(Al3+) cation (Positively charged ions) in soils, which also serves as an indicator of soil acidity. Agricultural practices and climate changes increase the amount of land affected by acidity and thus limit legume crop productivity. Worldwide, more than 1.5 Gha of acid soil limit agriculture production and as much as 25% of the earth’s crop lands are impacted by problems associated leguminous plants require a neutral or slightly acidic soil for growth, especially when depending on symbiotic nitrogen fixation. Out of several gases present in the atmosphere nitrogen share the major portion (about 71%) and is found in the di-nitrogen (an inert) form. It is a component of many bio-molecules required for the growth and development of all organisms. Most of the eukaryotes are capable of utilizing nitrogen directly from the environment; only a certain group of prokaryotes are genetically feasible to fix the atmospheric nitrogen into the biologically useful from like ammonia which is further utilized by eukaryotes. Rhizobia, a gram negative bacteria associates symbiotically with legume crop and are genetically feasible in reducing (fixing) atmospheric nitrogen for leguminous crop. The plant in turn provides the bacteria with organic compounds made by photosynthesis. The symbiotic factor like soil pH, temperature etc., which affects the survival of Rhizobia as well as the nodulation process and thus the nitrogen fixation. Soybean is scientifically known as Glycine max, are a species of legume that have become one of the most widely consumed foods in the world. Soybeans are rich in various bioactive plant compounds like Isoflavones, phytic acid, and saponins. Soybeans are high in protein, and are also a decent source of both carbs and fat. They are a rich source of various vitamins, minerals and beneficial plant compounds, such as Isoflavones. Proteomics is a large scale study of proteins. Two-dimensional electrophoresis (2-D electrophoresis) is a powerful and widely used method for the analysis of complex proteins mixtures extracted from cells, tissues, or other biological samples. This technique sort’s protein according to two independent properties in two discrete steps: the first-dimension step, isoelectric focusing (IEF), separates proteins according to their Isoelectric Point (pI); the second-dimension step, SDS-polyacrylamide gel electrophoresis (SDS-PAGE), separates proteins according to their molecular weight. Each spot on the resulting two-dimensional array corresponds to a single protein species in the sample. Thousands of different proteins can thus be separated, and information such as the protein pI, the apparent molecular weight, and the amount of each protein is obtained. The western blotting technique is a rapid and sensitive assay for detection and characterization of proteins. In this method labeled antibody against a particular protein is used to identify the desired protein, so it is a specific test. ELISA (Enzyme Immuno Sorbent Assay) is a technique used for detection of antigen or antibody. It is based on the formation of antigen-antibody complex, which is detected by chromogenic detection using enzyme. ELISA was performed at 405nm for pre immune sera, 1st and 3rd Bleed. HRP properties of the secondary antibodies were used to develop colour reactions. The ELISA results showed that 1st and 3rd bleed has a great titer for the antibodies produced. The ELISA also confirmed that the color reaction was obtained, thus showing the cross-reactivity to the protein specific to the acidic-soil regime in soybean crop. The purified antibodies can be used as bio-marker for the acidic soil regimes. The aim of present work is Protein characterization from Rhizobium isolates of Soybean [Glycine max (L.) Merrill] from acidic soils of State of Jharkhand by utilizing immunological approach. The present study was undertaken with the following broad objectives in mind: 1. ELISA analysis was performed for proteins of selected Rhizobium isolates from acidic-soils of State of Jharkhand, and 2. Generation of antibody profiles.ThesisItem Open Access Diversity analysis of endophytes of finger millet (Eleusinecoracana L. Gaertn.) using RAPD markers(Birsa Agricultural University, Ranchi, 2023) SWATI KUMARI; Anita PandeThe present study could show the closeness between the endophytes derived from one host versus those from a different host. The relatedness could help devise consortia of different endohytes for colonisation of other hosts once the role of the individual endophytes has been elucidated.ThesisItem Open Access Immunological characterization of various Rhizobium isolates from Pigeon pea (Cajanus Cajan (L.) Millsp.) isolated from acidic soils of the State of Jharkhand(Birsa Agricultural University, Ranchi, 2023) Aryan Raj; Himanshu DubeyWhen there is an accumulation of acid in the soil, soil acidity results. Almost 90% of the land in Jharkhand is covered with acidic soil. For optimal growth, most leguminous plants need neutral or slightly acidic soil, particularly when relying on symbiotic nitrogen fixation. Pigeon peas are legumes that improve soil fertility by biologically fixing nitrogen. According to reports, soil acidity is one of the major issues that affects the establishment of Rhizobia in the soil of Jharkhand and contributes around 40 kg N ha-1. Gram-negative, soil-dwelling bacteria are called rhizobium. Symbiosis is the relationship between the rhizobium, which delivers organic nitrogenous compound to the plant, and the plant, which produces compound through photosynthesis. An effective and popular technique for analyzing complex protein mixtures isolated from cells, tissues, or other biological materials is twodimensional electrophoresis (2-D electrophoresis). In two distinct processes, this method isolates proteins based on two independent properties: their isoelectric points (pI) in the first-dimension step of Isoelectric focusing (1EF), and their molecular weights in the second-dimension step of SDS polyacrylamide gel electrophoresis (SDS-PAGE). The resultant twodimensional array's spots each represent a distinct protein species from the sample. A list of experimental peptide masses, often known as "mass fingerprints," is produced by MALDI after it weighs peptides synthesized from trypsinized parent proteins. An ion's m/z ratio is calculated during MALDI-TOF analysis by timing how long it takes for it to go down the flight tube. A method for the specific histochemical demonstration of antibody in cells and parts of cells is described. It consists of carrying out a two stage immunological reaction on frozen sections of tissues: (a) allowing reaction between antibody in the tissue and dilute antigen applied in vitro, and (b) the detection of those areas where this antigen has been specifically absorbed by means of a precipitin reaction carried out with fluorescein-labelled antibody. The western blotting technique is a rapid and sensitive assay for detection and characterization of proteins. In this method labeled antibody against a particular protein is used to identify the desired protein, so it is a specific test. ELISA (Enzyme Immuno Sorbent Assay) is a technique used for detection of antigen or antibody. It is based on the formation of antigen-antibody complex, which is detected by chromogenic detection using enzyme. ELISA was performed at 405nm for pre immune sera, 1" and 3rd Bleed. HRP properties of the secondary antibodies were used to develop colour reactions. The ELISA results showed that 1st and 3rd bleed has a great titer for the antibodies produced. The ELISA also confirmed that the color reaction was obtained, thus showing the crossreactivity to the protein specific to the acidic-soil regime in soybean crop The aim of the work is immunological characterization of various Rhizobium isolates from Pigeon pea (Cajanus cajan (L.) Millsp.) isolated from acidic soils of the State of Jharkhand. The present study was undertaken with the following broad objectives in mind: 1. ELISA/Western Blot analysis was performed for proteins of selected Rhizobium isolates from acidic-soils of state of Jharkhand, 2. Generation of antibody profiles.ThesisItem Open Access Immunological Characterization of Rhizobium isoIlamtmesu fnroolmog cichaicl kcpheaar a(Ccitceerirz aartiioenti nouf mRh Liz.o) bisioulmat ed from acidic soils of the State of Jharkhand(Birsa Agricultural University, Ranchi, 2023) Poonam Priyanka TirkeySoil is a living, dynamic matrix that is vital to the terrestrial ecology. It is an essential resource for agricultural production and food security, as well as the preservation of most biological processes. Decomposition and nutrient cycling rely heavily on the operations of soil biota. As a result, the majority of microbial activity is restricted to the rhizosphere's "hot-spot," i.e., aggregates with accumulated organic matter. When there is a build-up of acid in the soil, it is known as soil acidity. Acid formation in soils is a natural process, and many soils in high-rainfall areas are acidic by nature. Acidification is a slow process, but agriculture hastens it. Plants that aren't tolerant of acidic conditions don't thrive in acidic soils, and productivity suffers as a result. Nitrogen is the most common deficient nutrient in many soils which are absorbed in the form of salts of nitrogen. Rhizobium are gram negative soil bacteria, motile, non-sporulating rods that fix atmospheric nitrogen i.e., convert atmospheric N2 into ammonia for their legume host plant once the symbiosis is established. Rhizobia are considered to be the most important nitrogen fixation agent in agriculture. N2 fixing systems are the symbiotic systems which play a significant role in improving the fertility and productivity of low- N soils. Chickpea (Cicer arietinum L.) is grain legumes grown mainly in areas with temperate and semiarid climate and about 400 millimetres of annual rainfall. It is characterized by a high content of protein, fat, vitamins, fibre, and a lower content of carbohydrates than flour of wheat. These are of two types “desi” and “kabuli” which are currently grown in India. Proteomics is a large scale study of proteins. Proteome techniques bring up new options to examine the complex functions of model plants and crop species at different levels. Proteomics include two-dimensional electrophoresis (2-D electrophoresis), separates proteins according to isoelectric points (pI) and SDS- Polyacrylamide gel electrophoresis (SDS-PAGE), and separates proteins according to molecular weight. These methods are used to analyse complex proteins mixture extracted from cells, tissues or other biological samples. Besides 2-D electrophoresis and SDS-PAGE, Western blot and ELISA are also used. ELISA is used to detect and measure antibodies, antigens, proteins, glycoproteins, and hormones, among other things. Antibodies and antigens are combined to create a measurable outcome in detection of these products. Western blot can use an antibody to specifically recognize its antigen. It separates different proteins in a sample using SDS-polyacrylamide gel electrophoresis (SDS-PAGE). In this study, soils from different locations were taken and landmark protein was identified. The two rabbits were employed through which antibodies were produced. ELISA analysis was done for antibodies detection. These antibodies can be used as biomarkers and the findings of this study provide a basis for further research on the use of Rhizobium inoculants in sustainable agriculture. The aim of present work is “Immunological characterization of Rhizobium isolates from chickpea (Cicer arietinum L.) isolated from acidic soils of the State Of Jharkhand”. The present study was undertaken with the following broad objectives in mind:ThesisItem Open Access Identification of drought specific cDNA in finger millet (Eleusine coracana L. Gaertn.) variety BM 3(Birsa Agricultural University, Ranchi, 2023) ANISHA KERKETTA; Anita PandeInformation generated in the present study will lead to greater understanding of the regulation of gene expression of finger millet under drought stress. It also shows that there may be some level of conservation or similarity between these different organisms at the genetic level when it comes to stress response. However, more research is needed to determine the significance of these findings and their potential impact on the biology of finger millet. It would be worth investigating further to determine the specific mechanisms by which these genetic similarities might be playing a role in the stress response of finger millet plants. This could potentially lead to the development of new techniques or technologies for improving crop resilience and yield under stress conditions.ThesisItem Open Access Diversity analysis of brinjal (Solanum melongena L.) genotypes using RAPD markers(Birsa Agricultural University, Ranchi, 2023) SUPRIYA BHARATI; Anita Pande; SUPRIYA BHARATIRAPD pattern can help to depict similarities and dissimilarities between different brinjal genotypes and support of phenotypic characterization. The present study shows that brinjal genotypes studied have a narrow genetic base. The use of RAPD markers for diversity analysis has been successfully demonstrated.ThesisItem Open Access Fingerprinting of Brinjal (Solanum melongena L.) genotypes using microsatellite markers(Birsa Agricultural University, Ranchi, 2023) BHARTI KUMARI; Anita PandeThe present study showed a very low degree of polymorphic present in the genotypes. This could be due to the selection of genotypes from a narrow base. Also, mostly compound microsatellites were used in present study the occurance of which may be infrequent. The use of simple repeats SSR and other markers may help in uncovering a larger number of polymorphic sites which these could then be used for generation of specific fringerprints.
