The presentation discuss about the present studies going on regarding biological nitrogen fixation in cereals. The material collected is recent one and includes all the potential studies and researches going around the world. the information given includes the pathways, the genetics, molecular mechanism, and results of various experiments and potential solutions. It describe all the possible ways and shows the future possibilities to achieve this dream.
Biological nitrogen fixation in cereal cropsAmandeep Kaur
The document discusses biological nitrogen fixation in cereal crops. It provides background on nitrogen and its importance for plant life. It describes how only certain prokaryotes and legumes can fix nitrogen. Efforts aim to transfer the nitrogen fixing ability of legumes to cereals by engineering cereal crops to recognize rhizobial signaling molecules and form nodule-like structures for nitrogen fixation. Key challenges include the oxygen sensitivity of the nitrogenase enzyme and competing demands for photosynthates. Transferring the nitrogen fixing genes or utilizing nitrogen fixing endophytes that associate with cereals are also approaches explored.
Lab.8 isolation of nitrogen fixer bacteriaHama Nabaz
Here are the answers to your questions:
1. The enzyme responsible for nitrogen fixation is nitrogenase. The level of O2 is regulated to obtain maximum nitrogenase activity through a process called microaerophily, where O2 concentration is kept low to protect the oxygen-sensitive nitrogenase enzyme.
2. Nitrogen fixation is important because atmospheric nitrogen gas (N2) is inert and unavailable to most organisms. Nitrogen fixation converts atmospheric N2 into bioavailable forms like ammonia (NH3) that can be used by plants and other organisms for growth. It replenishes soil nitrogen.
3. The Rhizobiaceae that form symbiotic relationships with legumes are more important than the Azotobacteraceae in
Biological nitrogen fixation (BNF) can be defined as the conversion of atmospheric dinitrogen (N2) to ammonia (NH3) under the combined action of biological and chemical activities
This document discusses Azotobacter, a genus of nitrogen-fixing bacteria that can be used as a biofertilizer. It describes the key species of Azotobacter, their identifying characteristics, and their benefits to agriculture. Azotobacter promotes plant growth by fixing atmospheric nitrogen and producing plant hormones. It also functions as a biocontrol agent by suppressing plant pathogens. The document outlines Azotobacter's mode of action in plants and provides examples of increased crop yields and quality from its use as an inoculant. It also discusses the maintenance, selection, and mass production methods for Azotobacter cultures.
Isolation of phosphate solubilizing bacteria (PSB) from soil Likhith KLIKHITHK1
A number of bacterial species provide beneficial effects to a plant and these are mostly present in rhizosphere and hence called rhizobacteria. This group of bacteria has been termed plant growth promoting rhizobacteria. Phosphorus is an essential element for plant development and growth making up about 0.2 % of plant dry weight. Plants acquire P from soil solution as phosphate anions. However, phosphate anions are extremely reactive and may be immobilized through precipitation with cations such as Ca 2+ , Mg 2+ , Fe 3+ and Al 3+. In these forms, P is highly insoluble and unavailable to plants. Different bacterial species has ability to solubilize insoluble inorganic phosphate compounds, such as tricalcium phosphate, di calcium phosphate, hydroxyapatite, and rock phosphate to soluble form, Hence theses bacteria's are referred to as phosphate solubilizing bacteria.
This document discusses biological nitrogen fixation, which is responsible for 65% of nitrogen used by humans through food. It occurs through nitrogen-fixing bacteria, which can be free-living like Azotobacter or symbiotic like Rhizobium that form nodules on legume roots. The bacteria contain the enzyme nitrogenase, which converts atmospheric nitrogen gas into ammonia in an oxygen-free environment within the nodules. The ammonia is then assimilated into amino acids and other biomolecules through a series of reactions.
Nitrogen fixation is the process by which atmospheric nitrogen is converted into nitrogen compounds that can be used by plants and other organisms. There are two main ways this occurs: biologically via certain bacteria, cyanobacteria, and symbiotic relationships with plants; and non-biologically through lightning and cosmic radiation. Biologically, nitrogen-fixing bacteria contain the enzyme nitrogenase, which converts nitrogen gas into ammonia. This process requires significant energy in the form of ATP. Nitrogen fixation is crucial for all life as it makes nitrogen available for use in important compounds like amino acids, proteins, and chlorophyll.
Here are the answers to the questions:
1. Nitrogen is present in the form of N2 gas in the atmosphere.
2. The enzyme nitrogenase is responsible for splitting the triple N≡N bond in nitrogen fixation.
3. The Haber process is used in chemical nitrogen fixation to convert nitrogen gas to ammonia at high pressure and temperature.
4. A nodulated symbiosis is the formation of nodules in leguminous plants by Rhizobium bacteria, for example Rhizobium and chickpea.
Biological nitrogen fixation in cereal cropsAmandeep Kaur
The document discusses biological nitrogen fixation in cereal crops. It provides background on nitrogen and its importance for plant life. It describes how only certain prokaryotes and legumes can fix nitrogen. Efforts aim to transfer the nitrogen fixing ability of legumes to cereals by engineering cereal crops to recognize rhizobial signaling molecules and form nodule-like structures for nitrogen fixation. Key challenges include the oxygen sensitivity of the nitrogenase enzyme and competing demands for photosynthates. Transferring the nitrogen fixing genes or utilizing nitrogen fixing endophytes that associate with cereals are also approaches explored.
Lab.8 isolation of nitrogen fixer bacteriaHama Nabaz
Here are the answers to your questions:
1. The enzyme responsible for nitrogen fixation is nitrogenase. The level of O2 is regulated to obtain maximum nitrogenase activity through a process called microaerophily, where O2 concentration is kept low to protect the oxygen-sensitive nitrogenase enzyme.
2. Nitrogen fixation is important because atmospheric nitrogen gas (N2) is inert and unavailable to most organisms. Nitrogen fixation converts atmospheric N2 into bioavailable forms like ammonia (NH3) that can be used by plants and other organisms for growth. It replenishes soil nitrogen.
3. The Rhizobiaceae that form symbiotic relationships with legumes are more important than the Azotobacteraceae in
Biological nitrogen fixation (BNF) can be defined as the conversion of atmospheric dinitrogen (N2) to ammonia (NH3) under the combined action of biological and chemical activities
This document discusses Azotobacter, a genus of nitrogen-fixing bacteria that can be used as a biofertilizer. It describes the key species of Azotobacter, their identifying characteristics, and their benefits to agriculture. Azotobacter promotes plant growth by fixing atmospheric nitrogen and producing plant hormones. It also functions as a biocontrol agent by suppressing plant pathogens. The document outlines Azotobacter's mode of action in plants and provides examples of increased crop yields and quality from its use as an inoculant. It also discusses the maintenance, selection, and mass production methods for Azotobacter cultures.
Isolation of phosphate solubilizing bacteria (PSB) from soil Likhith KLIKHITHK1
A number of bacterial species provide beneficial effects to a plant and these are mostly present in rhizosphere and hence called rhizobacteria. This group of bacteria has been termed plant growth promoting rhizobacteria. Phosphorus is an essential element for plant development and growth making up about 0.2 % of plant dry weight. Plants acquire P from soil solution as phosphate anions. However, phosphate anions are extremely reactive and may be immobilized through precipitation with cations such as Ca 2+ , Mg 2+ , Fe 3+ and Al 3+. In these forms, P is highly insoluble and unavailable to plants. Different bacterial species has ability to solubilize insoluble inorganic phosphate compounds, such as tricalcium phosphate, di calcium phosphate, hydroxyapatite, and rock phosphate to soluble form, Hence theses bacteria's are referred to as phosphate solubilizing bacteria.
This document discusses biological nitrogen fixation, which is responsible for 65% of nitrogen used by humans through food. It occurs through nitrogen-fixing bacteria, which can be free-living like Azotobacter or symbiotic like Rhizobium that form nodules on legume roots. The bacteria contain the enzyme nitrogenase, which converts atmospheric nitrogen gas into ammonia in an oxygen-free environment within the nodules. The ammonia is then assimilated into amino acids and other biomolecules through a series of reactions.
Nitrogen fixation is the process by which atmospheric nitrogen is converted into nitrogen compounds that can be used by plants and other organisms. There are two main ways this occurs: biologically via certain bacteria, cyanobacteria, and symbiotic relationships with plants; and non-biologically through lightning and cosmic radiation. Biologically, nitrogen-fixing bacteria contain the enzyme nitrogenase, which converts nitrogen gas into ammonia. This process requires significant energy in the form of ATP. Nitrogen fixation is crucial for all life as it makes nitrogen available for use in important compounds like amino acids, proteins, and chlorophyll.
Here are the answers to the questions:
1. Nitrogen is present in the form of N2 gas in the atmosphere.
2. The enzyme nitrogenase is responsible for splitting the triple N≡N bond in nitrogen fixation.
3. The Haber process is used in chemical nitrogen fixation to convert nitrogen gas to ammonia at high pressure and temperature.
4. A nodulated symbiosis is the formation of nodules in leguminous plants by Rhizobium bacteria, for example Rhizobium and chickpea.
This document discusses nitrogen fixation, which is the process of converting atmospheric nitrogen into ammonia. It is important because most organisms cannot use atmospheric nitrogen. The conversion is typically done by bacteria or cyanobacteria through one of two processes: symbiotic or non-symbiotic fixation. Symbiotic fixation often involves root nodules formed via infection by Rhizobia bacteria in legumes and some non-legumes. This process allows for nitrogen fixation to occur inside the plant.
Biology 12 - Solar Energy Converters - Section 7-2JEmmons
This document summarizes a chapter on photosynthesis from a biology textbook. It discusses how pigments like chlorophylls and carotenoids absorb different wavelengths of light during photosynthesis. Chlorophylls absorb violet, indigo, blue and red light most efficiently, while leaves appear green because chlorophyll reflects green light. The chapter then describes the two light reactions of photosynthesis - the noncyclic and cyclic electron pathways - and how they produce ATP and NADPH. It also explains how the proton gradient across the thylakoid membrane stores energy for the ATP synthase complex to generate ATP through chemiosmosis.
This document provides an overview of biofertilizers from an Indian perspective. It defines biofertilizers as microbial inoculants supported on carriers that are designed to improve soil fertility and provide growth promoters. The document discusses various types of biofertilizers including Rhizobium, Azotobacter, Azospirillum, cyanobacteria, and phosphate solubilizing microorganisms. It also covers India's history with biofertilizers, factors affecting their performance, application methods, production process, constraints to the industry's growth, and marketing challenges.
BIOFERTILIZERS: BENEFITS, PRODUCTION AND METHODS OF BIOFERTILIZER APPLICATION...Mahbubul Hassan
This document discusses biofertilizers, including their types and benefits. It defines biofertilizers as microorganisms used to supply nutrients to plants. The main types are nitrogen fixers like Rhizobium, and phosphate solubilizers like Pseudomonas. Biofertilizers can increase crop yields by 15-100% by fixing nitrogen, solubilizing phosphorus and potassium, and producing plant growth hormones. The document also describes procedures for producing and applying Rhizobium biofertilizer to legume seeds to maximize the benefits.
This document summarizes a seminar presentation about plant-microbe interactions given by Manisha Thakur. It discusses how plants constantly encounter biotic and abiotic stresses. Microbes that colonize plants can have pathogenic, symbiotic, or associative relationships. Specific examples provided include mutualistic relationships like rhizobia in root nodules, and types of pathogenic relationships such as necrotrophy and biotrophy. The document also discusses concepts like the rhizosphere and how root exudates influence microbial communities in and around plant roots and leaves.
This document discusses two types of nitrogen-fixing bacteria - Azotobacter and Azospirillum. Azotobacter is an aerobic, soil-dwelling bacteria that helps fix nitrogen. It is found in the rhizosphere of crops like rice and maize. Azospirillum is a microaerophilic, rod-shaped bacteria that colonizes plant surfaces and secretes plant hormones and nutrients to promote plant growth. Both bacteria play important roles in agriculture by increasing soil fertility and stimulating plant growth.
This document discusses various types of anaerobic respiration. It describes how anaerobic respiration works using electron acceptors other than oxygen, such as nitrates, sulfates, or carbon dioxide. It then examines different forms of anaerobic respiration in more detail, including denitrification, sulfate reduction, and sulfur disproportionation. Key enzymes and pathways involved in nitrate reduction, sulfate reduction, and other processes are outlined.
Avs role of plant growth promoting rhizobacteria in diseaseAMOL SHITOLE
This seminar discusses the role of plant growth promoting rhizobacteria (PGPR) in disease suppression and plant growth promotion. PGPR colonize plant roots and promote plant growth through mechanisms such as fixing atmospheric nitrogen, solubilizing mineral phosphates, producing phytohormones, antagonizing phytopathogenic microorganisms, and inducing systemic resistance in plants. The seminar outlines the definition of PGPR, common genera of PGPR including Pseudomonas and Bacillus, and the various mechanisms of action of PGPR such as nitrogen fixation, phosphate solubilization, phytohormone production, biocontrol activity, and induced systemic resistance. Experimental data is presented showing the effects of PGPR on nodulation,
Biological Nitrogen Fixation
Contents:
Introduction
Methods for measuring N2 fixation
1. Ntrogen balance method
2. Nitrogen difference method
3. Ureides method
4.〖𝟏𝟓〗_𝑵 isotope techniques
5. Acetylene reduction assay
6. Hydrogen evolution method
Introduction
N2 gas are found 78.084%on atmosphere of earth.
Nitrogen is an essential element for plant growth and development and a key issue of agriculture.
N2 are found in molecular N2 (𝑵 ≡ 𝑵) form in soil.
Dinitrogen is more stable, so we need of nitrogen fixation.
Most studies indicate that nitrogen fertilizers contribute to resolving the challenge the world is facing, feeding the human population.
The Green revolution was accompanied by an enormous increase in the application of nitrogen fertilizer.
Nitrogen fixation is a process by which nitrogen of the Earth's atmosphere is converted into ammonia (NH3), nitrogen salts or other molecules available to living organisms.
Biological Nitrogen Fixation(BNF) is known to be a sustain agriculture and increase soil fertility.
Research on microorganisms and plants able to fix nitrogen contributes largely to the production of bio fertilizers.
Thus it is important to ensure that BNF research and development will take into account the needs of farmers in the developing countries mainly.
Role of nitrogen in Plant
Sources of Nitrogen
Why measure 𝑵_𝟐 fixation?
Ecological consideration require an understanding of the relative contribution of 𝑵_𝟐 fixing components to the N-cycle.
Measurement of 𝑁_2 fixation enable an investigator to evaluate the ability of indigenous Rhizobium spp. to effectively nodulate newly introduced legumes.
Development of sustainable farming systems.
Understanding of the amount of 𝑵_𝟐fixed by legumes as influenced by soil management or cultural practices allows development of efficient agricultural and agroforesty production systems.
Nitrogen fixation is the process by which nitrogen in the atmosphere is converted into nitrogen compounds that can be used by plants. This can occur through non-biological means involving lightning and radiation, or through biological nitrogen fixation. Biological nitrogen fixation is performed by symbiotic and non-symbiotic bacteria and blue-green algae. Symbiotic nitrogen fixation occurs through root nodules formed by legumes via their symbiotic relationship with Rhizobia bacteria, and by some non-leguminous plants with actinomycetes. The bacteria in the nodules produce the nitrogenase enzyme which converts atmospheric nitrogen into ammonia that is used by the plant.
phyllosphere is a dynamic rapidly changing area surrounding the germinating seed. there are two categories of microbes one is positively enhancing and negatively reducing the plant yield
industrial applications of fungal proteasesrajani prabhu
- Proteases are enzymes that break down proteins and have many industrial uses. They are produced by animals, plants, and microbes. Microbial sources such as fungi are preferred for large-scale production due to their fast growth.
- Fungal proteases exhibit high diversity, broad substrate specificity, and stability under extreme conditions. Important fungal genera producing commercial proteases include Aspergillus, Beauveria, and Acremonium. These proteases have applications in food processing, textiles, detergents, leather processing, and more.
- Proteases are classified based on factors such as their site of action, pH optimum, and catalytic mechanism. Major classes include serine, aspart
This document discusses the nitrogen cycle and nitrogen fixation. It describes how nitrogen exists in the atmosphere as N2 gas which requires a lot of energy to convert to other forms usable by organisms. Some key points made are:
- Certain bacteria and archaea are able to fix nitrogen through the enzyme nitrogenase. This can occur freely in the environment or through symbiotic relationships with plants.
- Legumes form nodules on their roots housing rhizobia bacteria which are able to fix atmospheric nitrogen. The nodules provide protection from oxygen for the oxygen-sensitive nitrogenase enzyme.
- Fixed nitrogen is used to synthesize amino acids within plants through reductive amination or transamination reactions. Excess nitrogen is
Biomining and bioleaching use microorganisms like Thiobacillus ferrooxidans to extract metals from ores and mine tailings. These microbes facilitate metal extraction by oxidizing metals or the minerals containing them, making the metals soluble so they can be recovered. Key applications include extracting copper, gold, and uranium, as well as remediating acid mine drainage. As high-grade surface deposits diminish, biomining will become increasingly important for recovering metals from lower-grade ores in a more sustainable and cost-effective manner than conventional mining and extraction methods.
This document discusses the biochemical composition and biodegradation of organic matter in soils. It describes the various components of soil organic matter including nitrogenous and non-nitrogenous organic compounds. It explains the roles of enzymes and microbes like fungi, bacteria, and actinomycetes in decomposing different organic compounds such as proteins, cellulose, hemicellulose, starch, and lignin. Finally, it outlines several factors that affect the rate of organic matter decomposition in soils like temperature, moisture, nutrients, pH, texture and toxic elements.
This document provides information on biofertilizers. It discusses that biofertilizers are nutrient inputs of biological origin that aid plant growth through microbiological processes. Important microorganisms used as biofertilizers include nitrogen-fixing bacteria, fungi, cyanobacteria, and ferns. These microorganisms can fix nitrogen either symbiotically, by forming relationships with plants, or non-symbiotically as free-living organisms. The document also describes methods for mass cultivating and applying different types of biofertilizer microorganisms, such as Rhizobium bacteria and blue-green algae, to benefit agricultural crops.
Microbial biomass in soil, measurement by chloroform fumigation incubation method, limits of measurement of microbial biomass, why microbes are important in the soil, why microbial biomass is important in the soil
Non-symbiotic nitrogen fixation refers to nitrogen fixation by microorganisms that live independently in soil and aquatic environments. Key microorganisms that can fix nitrogen this way include species from the genera Azotobacter, Clostridium, Klebsiella, cyanobacteria, and some fungi. Azotobacter is a common soil bacterium that can fix significant amounts of nitrogen aerobically using the nitrogenase enzyme. Cyanobacteria also fix nitrogen, with some genera like Nostoc forming specialized cells called heterocysts that protect the oxygen-sensitive nitrogenase enzyme.
Viruses are being explored as potential biopesticides to control insect pests. The major viruses investigated are baculoviruses, which primarily infect lepidopteran insects. Baculoviruses are classified as nucleopolyhedroviruses (NPVs) or granuloviruses (GVs) depending on how their virions are occluded. NPVs occlude virions in large polyhedral bodies, while GVs occlude individual virions. These viruses replicate in the nucleus or cytoplasm of infected insects and cause symptoms like discoloration, lethargy, and death. Large-scale production can be done in vivo by applying the virus to host insects or in vitro by infecting insect
This document discusses non-symbiotic nitrogen fixation by microorganisms. It explains that nitrogen is essential for plants and must be fixed from its atmospheric form (N2) into nitrogen salts that plants can absorb. This is accomplished through nitrogen fixation, where microorganisms like bacteria and cyanobacteria are able to convert N2 into ammonia using the nitrogenase enzyme. The document focuses on non-symbiotic nitrogen fixation, which is performed by free-living microorganisms like aerobic and anaerobic bacteria and blue-green algae, as opposed to symbiotic fixation that involves nodule formation.
1) Soil microbes play an important role in global carbon and nitrogen cycles by driving processes like organic matter decomposition, nitrogen fixation, and methane/nitrous oxide emissions.
2) Changing environmental conditions due to climate change, like increased temperature and altered precipitation, can impact soil microbial communities and gene expression.
3) Horizontal gene transfer between soil microbes may be a natural adaptation strategy to environmental changes, allowing microbes to acquire new genes that help them survive. Studying this process of natural transformation could provide insights into molecular-level climate change adaptation.
This document discusses nitrogen fixation, which is the process of converting atmospheric nitrogen into ammonia. It is important because most organisms cannot use atmospheric nitrogen. The conversion is typically done by bacteria or cyanobacteria through one of two processes: symbiotic or non-symbiotic fixation. Symbiotic fixation often involves root nodules formed via infection by Rhizobia bacteria in legumes and some non-legumes. This process allows for nitrogen fixation to occur inside the plant.
Biology 12 - Solar Energy Converters - Section 7-2JEmmons
This document summarizes a chapter on photosynthesis from a biology textbook. It discusses how pigments like chlorophylls and carotenoids absorb different wavelengths of light during photosynthesis. Chlorophylls absorb violet, indigo, blue and red light most efficiently, while leaves appear green because chlorophyll reflects green light. The chapter then describes the two light reactions of photosynthesis - the noncyclic and cyclic electron pathways - and how they produce ATP and NADPH. It also explains how the proton gradient across the thylakoid membrane stores energy for the ATP synthase complex to generate ATP through chemiosmosis.
This document provides an overview of biofertilizers from an Indian perspective. It defines biofertilizers as microbial inoculants supported on carriers that are designed to improve soil fertility and provide growth promoters. The document discusses various types of biofertilizers including Rhizobium, Azotobacter, Azospirillum, cyanobacteria, and phosphate solubilizing microorganisms. It also covers India's history with biofertilizers, factors affecting their performance, application methods, production process, constraints to the industry's growth, and marketing challenges.
BIOFERTILIZERS: BENEFITS, PRODUCTION AND METHODS OF BIOFERTILIZER APPLICATION...Mahbubul Hassan
This document discusses biofertilizers, including their types and benefits. It defines biofertilizers as microorganisms used to supply nutrients to plants. The main types are nitrogen fixers like Rhizobium, and phosphate solubilizers like Pseudomonas. Biofertilizers can increase crop yields by 15-100% by fixing nitrogen, solubilizing phosphorus and potassium, and producing plant growth hormones. The document also describes procedures for producing and applying Rhizobium biofertilizer to legume seeds to maximize the benefits.
This document summarizes a seminar presentation about plant-microbe interactions given by Manisha Thakur. It discusses how plants constantly encounter biotic and abiotic stresses. Microbes that colonize plants can have pathogenic, symbiotic, or associative relationships. Specific examples provided include mutualistic relationships like rhizobia in root nodules, and types of pathogenic relationships such as necrotrophy and biotrophy. The document also discusses concepts like the rhizosphere and how root exudates influence microbial communities in and around plant roots and leaves.
This document discusses two types of nitrogen-fixing bacteria - Azotobacter and Azospirillum. Azotobacter is an aerobic, soil-dwelling bacteria that helps fix nitrogen. It is found in the rhizosphere of crops like rice and maize. Azospirillum is a microaerophilic, rod-shaped bacteria that colonizes plant surfaces and secretes plant hormones and nutrients to promote plant growth. Both bacteria play important roles in agriculture by increasing soil fertility and stimulating plant growth.
This document discusses various types of anaerobic respiration. It describes how anaerobic respiration works using electron acceptors other than oxygen, such as nitrates, sulfates, or carbon dioxide. It then examines different forms of anaerobic respiration in more detail, including denitrification, sulfate reduction, and sulfur disproportionation. Key enzymes and pathways involved in nitrate reduction, sulfate reduction, and other processes are outlined.
Avs role of plant growth promoting rhizobacteria in diseaseAMOL SHITOLE
This seminar discusses the role of plant growth promoting rhizobacteria (PGPR) in disease suppression and plant growth promotion. PGPR colonize plant roots and promote plant growth through mechanisms such as fixing atmospheric nitrogen, solubilizing mineral phosphates, producing phytohormones, antagonizing phytopathogenic microorganisms, and inducing systemic resistance in plants. The seminar outlines the definition of PGPR, common genera of PGPR including Pseudomonas and Bacillus, and the various mechanisms of action of PGPR such as nitrogen fixation, phosphate solubilization, phytohormone production, biocontrol activity, and induced systemic resistance. Experimental data is presented showing the effects of PGPR on nodulation,
Biological Nitrogen Fixation
Contents:
Introduction
Methods for measuring N2 fixation
1. Ntrogen balance method
2. Nitrogen difference method
3. Ureides method
4.〖𝟏𝟓〗_𝑵 isotope techniques
5. Acetylene reduction assay
6. Hydrogen evolution method
Introduction
N2 gas are found 78.084%on atmosphere of earth.
Nitrogen is an essential element for plant growth and development and a key issue of agriculture.
N2 are found in molecular N2 (𝑵 ≡ 𝑵) form in soil.
Dinitrogen is more stable, so we need of nitrogen fixation.
Most studies indicate that nitrogen fertilizers contribute to resolving the challenge the world is facing, feeding the human population.
The Green revolution was accompanied by an enormous increase in the application of nitrogen fertilizer.
Nitrogen fixation is a process by which nitrogen of the Earth's atmosphere is converted into ammonia (NH3), nitrogen salts or other molecules available to living organisms.
Biological Nitrogen Fixation(BNF) is known to be a sustain agriculture and increase soil fertility.
Research on microorganisms and plants able to fix nitrogen contributes largely to the production of bio fertilizers.
Thus it is important to ensure that BNF research and development will take into account the needs of farmers in the developing countries mainly.
Role of nitrogen in Plant
Sources of Nitrogen
Why measure 𝑵_𝟐 fixation?
Ecological consideration require an understanding of the relative contribution of 𝑵_𝟐 fixing components to the N-cycle.
Measurement of 𝑁_2 fixation enable an investigator to evaluate the ability of indigenous Rhizobium spp. to effectively nodulate newly introduced legumes.
Development of sustainable farming systems.
Understanding of the amount of 𝑵_𝟐fixed by legumes as influenced by soil management or cultural practices allows development of efficient agricultural and agroforesty production systems.
Nitrogen fixation is the process by which nitrogen in the atmosphere is converted into nitrogen compounds that can be used by plants. This can occur through non-biological means involving lightning and radiation, or through biological nitrogen fixation. Biological nitrogen fixation is performed by symbiotic and non-symbiotic bacteria and blue-green algae. Symbiotic nitrogen fixation occurs through root nodules formed by legumes via their symbiotic relationship with Rhizobia bacteria, and by some non-leguminous plants with actinomycetes. The bacteria in the nodules produce the nitrogenase enzyme which converts atmospheric nitrogen into ammonia that is used by the plant.
phyllosphere is a dynamic rapidly changing area surrounding the germinating seed. there are two categories of microbes one is positively enhancing and negatively reducing the plant yield
industrial applications of fungal proteasesrajani prabhu
- Proteases are enzymes that break down proteins and have many industrial uses. They are produced by animals, plants, and microbes. Microbial sources such as fungi are preferred for large-scale production due to their fast growth.
- Fungal proteases exhibit high diversity, broad substrate specificity, and stability under extreme conditions. Important fungal genera producing commercial proteases include Aspergillus, Beauveria, and Acremonium. These proteases have applications in food processing, textiles, detergents, leather processing, and more.
- Proteases are classified based on factors such as their site of action, pH optimum, and catalytic mechanism. Major classes include serine, aspart
This document discusses the nitrogen cycle and nitrogen fixation. It describes how nitrogen exists in the atmosphere as N2 gas which requires a lot of energy to convert to other forms usable by organisms. Some key points made are:
- Certain bacteria and archaea are able to fix nitrogen through the enzyme nitrogenase. This can occur freely in the environment or through symbiotic relationships with plants.
- Legumes form nodules on their roots housing rhizobia bacteria which are able to fix atmospheric nitrogen. The nodules provide protection from oxygen for the oxygen-sensitive nitrogenase enzyme.
- Fixed nitrogen is used to synthesize amino acids within plants through reductive amination or transamination reactions. Excess nitrogen is
Biomining and bioleaching use microorganisms like Thiobacillus ferrooxidans to extract metals from ores and mine tailings. These microbes facilitate metal extraction by oxidizing metals or the minerals containing them, making the metals soluble so they can be recovered. Key applications include extracting copper, gold, and uranium, as well as remediating acid mine drainage. As high-grade surface deposits diminish, biomining will become increasingly important for recovering metals from lower-grade ores in a more sustainable and cost-effective manner than conventional mining and extraction methods.
This document discusses the biochemical composition and biodegradation of organic matter in soils. It describes the various components of soil organic matter including nitrogenous and non-nitrogenous organic compounds. It explains the roles of enzymes and microbes like fungi, bacteria, and actinomycetes in decomposing different organic compounds such as proteins, cellulose, hemicellulose, starch, and lignin. Finally, it outlines several factors that affect the rate of organic matter decomposition in soils like temperature, moisture, nutrients, pH, texture and toxic elements.
This document provides information on biofertilizers. It discusses that biofertilizers are nutrient inputs of biological origin that aid plant growth through microbiological processes. Important microorganisms used as biofertilizers include nitrogen-fixing bacteria, fungi, cyanobacteria, and ferns. These microorganisms can fix nitrogen either symbiotically, by forming relationships with plants, or non-symbiotically as free-living organisms. The document also describes methods for mass cultivating and applying different types of biofertilizer microorganisms, such as Rhizobium bacteria and blue-green algae, to benefit agricultural crops.
Microbial biomass in soil, measurement by chloroform fumigation incubation method, limits of measurement of microbial biomass, why microbes are important in the soil, why microbial biomass is important in the soil
Non-symbiotic nitrogen fixation refers to nitrogen fixation by microorganisms that live independently in soil and aquatic environments. Key microorganisms that can fix nitrogen this way include species from the genera Azotobacter, Clostridium, Klebsiella, cyanobacteria, and some fungi. Azotobacter is a common soil bacterium that can fix significant amounts of nitrogen aerobically using the nitrogenase enzyme. Cyanobacteria also fix nitrogen, with some genera like Nostoc forming specialized cells called heterocysts that protect the oxygen-sensitive nitrogenase enzyme.
Viruses are being explored as potential biopesticides to control insect pests. The major viruses investigated are baculoviruses, which primarily infect lepidopteran insects. Baculoviruses are classified as nucleopolyhedroviruses (NPVs) or granuloviruses (GVs) depending on how their virions are occluded. NPVs occlude virions in large polyhedral bodies, while GVs occlude individual virions. These viruses replicate in the nucleus or cytoplasm of infected insects and cause symptoms like discoloration, lethargy, and death. Large-scale production can be done in vivo by applying the virus to host insects or in vitro by infecting insect
This document discusses non-symbiotic nitrogen fixation by microorganisms. It explains that nitrogen is essential for plants and must be fixed from its atmospheric form (N2) into nitrogen salts that plants can absorb. This is accomplished through nitrogen fixation, where microorganisms like bacteria and cyanobacteria are able to convert N2 into ammonia using the nitrogenase enzyme. The document focuses on non-symbiotic nitrogen fixation, which is performed by free-living microorganisms like aerobic and anaerobic bacteria and blue-green algae, as opposed to symbiotic fixation that involves nodule formation.
1) Soil microbes play an important role in global carbon and nitrogen cycles by driving processes like organic matter decomposition, nitrogen fixation, and methane/nitrous oxide emissions.
2) Changing environmental conditions due to climate change, like increased temperature and altered precipitation, can impact soil microbial communities and gene expression.
3) Horizontal gene transfer between soil microbes may be a natural adaptation strategy to environmental changes, allowing microbes to acquire new genes that help them survive. Studying this process of natural transformation could provide insights into molecular-level climate change adaptation.
This document summarizes the key outcomes, background, significance, and approach of a study that investigated the effects of glycerol thermal processing (GTP) on lignin structure through NMR analysis. The main outcomes were that GTP effectively breaks bonds within lignin, generating smaller molecular weight lignin fragments that are thermally stable up to temperatures over 290°C. The background discusses utilizing lignin for specialty products. The significance is that GTP generates lignin suitable for use as a thermoplastic co-product. The approach involved comparing the properties of GTP lignin to lignin from other processes through structural analysis.
Research from a bacterium bacillus subtilis b 3157 by fabAlexander Decker
This document discusses research into the biosynthetic pathways that produce 2H-labeled inosine in the bacterium Bacillus subtilis B-3157. The bacterium was grown in heavy water medium containing a hydrolysate of deuterated biomass as a source of 2H-labeled substrates. Isolation and analysis of the produced 2H-labeled inosine found incorporation of 5 deuterium atoms, with 3 in the ribose residue and 2 in the hypoxanthine residue. The non-exchangeable deuterium atoms in ribose originated from HMP shunt reactions, while the atoms in hypoxanthine came from [2H]amino acids in the growth medium.
This document discusses microbial fuel cells (MFCs) which generate electricity through the catalytic activity of microorganisms. MFCs convert the chemical energy in organic matter like biomass or biofuel into electricity. They have several advantages like satisfying increasing energy demand, decreasing dependence on fossil fuels, and generating clean energy. MFCs use microbes like bacteria to catalyze the oxidation of an organic compound at the anode and reduction of oxygen at the cathode. Various wastewaters have been used as substrates to power MFCs. Field applications of MFCs include powering sensors for oceanic monitoring, tsunami prediction, and groundwater nitrate detection.
This document discusses microbial fuel cells (MFCs) which are bio-electrochemical systems that convert the chemical energy in organic matter into electricity through catalytic microorganisms. MFCs have several applications including powering low-power sensors for oceanographic monitoring and environmental applications. The document outlines the basic process of an MFC where microbes oxidize a substrate like glucose at the anode and electrons are transferred to the cathode via a membrane or mediator. Various types of wastewater can be used as substrates to power MFCs. Common microbes used in MFCs include Geobacter and Shewanella species.
Presentation on genetics of nitrogen fixation by Tahura MariyamTahura Mariyam Ansari
this presentation is about what is the genetics involvement in nitrogen fixation i.e which gene is responsible etc....
the contents include Genetics of N2 fixing microorganisms, Bacterial Nodulation Genes and Regulation of nod Gene Expression, Nif Genes and their Regulation in K. Pneumoniae & Cyanobacteria, Nitrogen fixation mechanism
Nitrogenase Types, Structure and Function, Alternative nitrogenase, Substrate for Nitrogenase, Electron proteins and Hydrogen evolution
Role of Biofortification in Combating Zinc & Iron DeficiencyHimanshu Pandey
Biofortification stands as a pivotal strategy in combating zinc and iron deficiencies, particularly in regions grappling with limited access to diverse diets or nutritional supplements. Large scale occurrences of zinc and iron deficiencies in the Indian population are associated with production of staple food grains low in these nutrients and are recognized as the key factors behind human malnutrition. Biofortified crops not only enhance the nutrient content of staple foods but also integrate vital minerals directly into local food systems, increasing accessibility, especially in remote or rural areas. Moreover, the cultural acceptance of biofortified crops is often high, as they are developed through traditional breeding methods and closely resemble local varieties in taste and appearance. This fosters their adoption by communities, further amplifying their impact. Importantly, biofortification is a cost-effective approach that leverages existing agricultural infrastructure, making it feasible for large-scale implementation.
By providing sustainable sources of zinc and iron, biofortified crops contribute to improving health outcomes, particularly among vulnerable populations such as children and pregnant women. Ultimately, by addressing hidden hunger and bolstering nutritional intake, biofortification plays a vital role in promoting public health and combating malnutrition globally. Biofortified crops offer a sustainable solution to the problem of nutrient deficiencies. Through targeted breeding efforts, crop varieties with elevated levels of zinc and iron can be developed, ensuring that these essential minerals are naturally present in staple foods like rice, wheat, maize, and beans. This approach bypasses the need for external interventions such as nutritional supplements or fortified foods, which may not always be readily available or affordable, especially in rural or underserved areas.
Indian agriculture feels the pain of fatigue of green revolution.
In the past 50 years, the fertilizer consumption exponentially increased from 0.5 (1960’s) to 24 million tonnes (2013) that commensurate with four-fold increase in food grain output (254 million tonnes) In order to achieve a target of 300 million tonnes of food grains and to feed the burgeoning population of 1.4 billion in 2025, the country will require 45 million tonnes of nutrients as against a current consumption level of 23 million tonnes. The sustainable agriculture and precision farming both are the urgent issues and hence the suitable agro-technological interventions are essential (e.g., nano and biotechnology) for ensuring the safety and sustainability of relevant production system.
This document describes the development of a genetically encoded fluorescent biosensor for 2-oxoglutarate (2OG) based on fluorescence resonance energy transfer (FRET). The researchers constructed the biosensor by fusing yellow fluorescent protein (YFP), the 2OG-binding GAF domain of the NifA protein, and cyan fluorescent protein (CFP) between restriction sites in a plasmid vector. They tested the biosensor in vitro and found it responded to 2OG concentrations within the physiological range observed in E. coli cells. They also optimized the peptide linker between domains. In vivo, the biosensor detected changes in 2OG levels in E. coli cells in response to different carbon sources added. The biosensor could image
Maize (Zea mays L.) and wheat [Triticum aestivum (L.) emend. Fiori & Paol] is the third and second most important cereal crop of India, respectively. Maize–wheat system is the third dominant cropping system of India covering 1.8 mha with 2.3% contribution in food grain production (Jat et al., 2013).
Interactions between nutrients in plants occur when the supply of one nutrient affects the absorption, distribution and functions of another nutrient. Generally P and Zn interact negatively, which depends upon a number of physico-chemical properties of soil. Antagonistic P×Zn interaction has been subject of intensive research in several countries and has been thoroughly reviewed. Although some positive interactions of P and Zn are also reported (Shivay, 2013).
The maximum available P and Zn content in the soil was recorded with super-optimal dose (150% NPK) and optimal dose (100% NPK) along with Zn, respectively (Verma et al., 2012). Zinc and P application has antagonistic effect on each other with respect to their concentration and absorption by wheat and maize (Verma and Minhas, 1987). The three Bacillus aryabhattai strains (MDSR7, MDSR11 and MDSR14) were consistent in enhancement of root and shoot dry weight and zinc uptake in wheat (Ramesh et al., 2014).
Management of P×Zn interaction is a challenging task in the era of sustainable food and nutritional security. Use of efficient varieties and application of inorganic P and Zn fertilizer in conjunction with bio-inoculants can increase the crop yield and efficiency of added fertilizers to save precious input.
Agronomic-fortification is one such approach that involves the application of foliar fertilizers or combined soil
and foliar fertilizers, intercropping with pulse and crop rotation, which is a highly effective and practical way to
maximize the absorption and accumulation of micronutrients in the grain. It is also recognized as one of the cheapest
ways to reduce mineral deficiency in the human diet.
This study assessed whether introducing a legume pasture in a subtropical cereal cropping system can reduce synthetic nitrogen (N) inputs and nitrous oxide (N2O) emissions. The study compared N2O emissions and yields in a sorghum crop following either a legume pasture (alfalfa and sulla) or grass pasture (rhodes grass and wheat) rotation under two N fertilization rates. N2O emissions were monitored from crop planting to final seedbed preparation using an automated system. Preliminary results showed that the legume pasture supplied enough N to support crop growth while low carbon residues limited denitrification and N2O emissions compared to the grass pasture. Introducing a leg
Organic farming with special reference to vermicultureTakeleZike1
This presentation delves into the principles and practices of organic farming, with a particular focus on the innovative technique of vermiculture. Organic farming represents a sustainable approach to agriculture that emphasizes the use of natural inputs and biological processes to enhance soil fertility, conserve resources, and minimize environmental impact. Within this context, vermiculture, or the use of earthworms to process organic waste and create nutrient-rich vermicompost, emerges as a powerful tool for organic farmers.
Throughout the presentation, key aspects of organic farming are explored, including soil health management, crop rotation, companion planting, and natural pest control methods. The role of vermiculture in organic farming systems is examined in depth, highlighting its benefits in improving soil structure, increasing microbial activity, and supplying essential nutrients to plants. Practical guidance is provided on setting up and managing a vermiculture system, from selecting suitable earthworm species to optimizing environmental conditions for composting.
Case studies and examples illustrate the real-world applications of organic farming and vermiculture, showcasing successful initiatives and their positive impact on agricultural sustainability, biodiversity, and food security. By promoting a holistic and ecologically sound approach to agriculture, this presentation aims to inspire farmers, researchers, and policymakers to embrace organic farming practices, with vermiculture as a valuable component in achieving long-term agricultural resilience and environmental stewardship.
1. Di-nitrogen was produced both abiotically and in the presence of live and dead fungi, with no evidence that N2O consumption was required for N2 production.
2. Isotope pairing experiments indicated the N2 was produced abiotically by the combination of glutamine nitrogen and nitrite nitrogen.
3. Di-nitrogen was produced abiotically under both anaerobic and aerobic conditions, calling into question the assumptions that anaerobic conditions and N2O production are required for N2 formation.
This document describes an efficient synthetic route to dihydropipercide and pipercide. Key steps include the preparation of methyl 6-oxohexanoate from cyclohexanone via ozonolysis and methanolysis. A Wittig olefination reaction is used to couple methyl 6-oxohexanoate with a phosphonium salt derived from piperonyl bromide. Phenythio radical-induced olefin isomerization is then used to obtain the desired (E)-olefin geometry. Further elaboration allows construction of the targets via a Horner-Emmons reaction and amidation. The described methods provide practical gram-scale syntheses of these biologically active dieneamide natural products.
Agriculture met the challenge of feeding the world’s poor by the Green Revolution with the help of high yielding varieties (HYV), high fertilizer application. This high fertilizer application increased the world food grain production as well as micro nutrient deficiencies in the soil decade to decade. in 1950 only Nitrogen is deficient in soil but due to green revolution, higher fertilizer application leads to micro nutrient deficiencies in soil (Fig.1). Iron, zinc and Vitamin A deficiencies in human nutrition are widespread in developing countries. About 2 billion people suffer globally from anaemia due to Fe deficiency, more than one-third of the world’s population suffers from Zn deficiency and estimated to be responsible for approximately 4% of the worldwide burden of morbidity and mortality in under 5-year children.
Bio-fortification entails the development of micronutrient-dense food crops (Nestel et al., 2006). Plant breeding strategies hold great promise in this process because of its enormous potential to improve dietary quality. Well-known examples of bio-fortification for fighting micronutrient malnutrition are golden rice and breeding of low phytate legumes and grains (Beyer et al., 2006). Application of fertilizers to soil and/or foliar to improving grain nutrient concentration and the potential of nutrient containing fertilizers for increasing nutrient concentration of cereal grains. Increasing the Zn and Fe concentration of food crop plants, resulting in better crop production and improved human health is an important global challenge. Among micronutrients, Zn and Fe deficiency are occurring in both crops and humans. Zinc deficiency is currently listed as a major risk factor for human health and cause of death globally.
In view of globally widespread deficiencies of micronutrients in humans, bio-fortification of food crops with micronutrients through agricultural approaches is a sustainable widely applied strategy. Agronomic bio-fortification (e.g., fertilizer applications) and plant breeding (e.g., genetic bio-fortification and transgenic breeding) represent complementary and cost-effective solution to alleviate malnutrition. Bio-fortified varieties assume great significance to achieve nutritional security of the country.
Micronutrient malnutrition Causes….
• More severe illness
• More infant and maternal deaths
• Lower cognitive development
• Stunted growth
• Lower work productivity and ultimately - Lower GDP.
• Higher population growth rates.
Malnutrition Problem
• 800 million people go to bed hungry
• 250 million children are malnourished
• 400 million people have vitamin A deficiency
• 100 million young children suffer from vitamin A deficiency
• 3 million children die as a result of vitamin A deficiency
Estimating the Biodegradation Kinetics by Mixed Culture Degrading Pyrene (Pyr)AZOJETE UNIMAID
This document discusses the biodegradation kinetics of pyrene (Pyr) degradation by a mixed culture. Experiments were conducted to determine the optimal temperature, pH, and Pyr concentrations for degradation. Kinetics experiments were then carried out at 30°C and pH 7 using initial Pyr concentrations ranging from 10-700 ppm. The results showed Pyr concentrations between 100-700 ppm inhibited the mixed culture. Concentrations between 10-100 ppm did not inhibit growth. A first-order rate constant model best described the degradation kinetics, with the highest rate (0.0487 mg/Lh) at 20 ppm Pyr. On average, the mixed culture could degrade over 0.0696 p
PULSE CROPS FOR SUSTAINABLE PRODUCTION INTENSIFICATIONExternalEvents
http://www.fao.org/globalsoilpartnership/en/
This presentation was presentaed during the seminar Soils & Pulses: symbiosis for life that took place at FAO HQ on 19 Apr 2016. it was made by Paola De Santis and it presents the using op pulses diversity.
Semelhante a A biotechnology dream nitrogen fixing cereal crops by Deepak Sharma (20)
When I was asked to give a companion lecture in support of ‘The Philosophy of Science’ (https://shorturl.at/4pUXz) I decided not to walk through the detail of the many methodologies in order of use. Instead, I chose to employ a long standing, and ongoing, scientific development as an exemplar. And so, I chose the ever evolving story of Thermodynamics as a scientific investigation at its best.
Conducted over a period of >200 years, Thermodynamics R&D, and application, benefitted from the highest levels of professionalism, collaboration, and technical thoroughness. New layers of application, methodology, and practice were made possible by the progressive advance of technology. In turn, this has seen measurement and modelling accuracy continually improved at a micro and macro level.
Perhaps most importantly, Thermodynamics rapidly became a primary tool in the advance of applied science/engineering/technology, spanning micro-tech, to aerospace and cosmology. I can think of no better a story to illustrate the breadth of scientific methodologies and applications at their best.
EWOCS-I: The catalog of X-ray sources in Westerlund 1 from the Extended Weste...Sérgio Sacani
Context. With a mass exceeding several 104 M⊙ and a rich and dense population of massive stars, supermassive young star clusters
represent the most massive star-forming environment that is dominated by the feedback from massive stars and gravitational interactions
among stars.
Aims. In this paper we present the Extended Westerlund 1 and 2 Open Clusters Survey (EWOCS) project, which aims to investigate
the influence of the starburst environment on the formation of stars and planets, and on the evolution of both low and high mass stars.
The primary targets of this project are Westerlund 1 and 2, the closest supermassive star clusters to the Sun.
Methods. The project is based primarily on recent observations conducted with the Chandra and JWST observatories. Specifically,
the Chandra survey of Westerlund 1 consists of 36 new ACIS-I observations, nearly co-pointed, for a total exposure time of 1 Msec.
Additionally, we included 8 archival Chandra/ACIS-S observations. This paper presents the resulting catalog of X-ray sources within
and around Westerlund 1. Sources were detected by combining various existing methods, and photon extraction and source validation
were carried out using the ACIS-Extract software.
Results. The EWOCS X-ray catalog comprises 5963 validated sources out of the 9420 initially provided to ACIS-Extract, reaching a
photon flux threshold of approximately 2 × 10−8 photons cm−2
s
−1
. The X-ray sources exhibit a highly concentrated spatial distribution,
with 1075 sources located within the central 1 arcmin. We have successfully detected X-ray emissions from 126 out of the 166 known
massive stars of the cluster, and we have collected over 71 000 photons from the magnetar CXO J164710.20-455217.
Immersive Learning That Works: Research Grounding and Paths ForwardLeonel Morgado
We will metaverse into the essence of immersive learning, into its three dimensions and conceptual models. This approach encompasses elements from teaching methodologies to social involvement, through organizational concerns and technologies. Challenging the perception of learning as knowledge transfer, we introduce a 'Uses, Practices & Strategies' model operationalized by the 'Immersive Learning Brain' and ‘Immersion Cube’ frameworks. This approach offers a comprehensive guide through the intricacies of immersive educational experiences and spotlighting research frontiers, along the immersion dimensions of system, narrative, and agency. Our discourse extends to stakeholders beyond the academic sphere, addressing the interests of technologists, instructional designers, and policymakers. We span various contexts, from formal education to organizational transformation to the new horizon of an AI-pervasive society. This keynote aims to unite the iLRN community in a collaborative journey towards a future where immersive learning research and practice coalesce, paving the way for innovative educational research and practice landscapes.
Describing and Interpreting an Immersive Learning Case with the Immersion Cub...Leonel Morgado
Current descriptions of immersive learning cases are often difficult or impossible to compare. This is due to a myriad of different options on what details to include, which aspects are relevant, and on the descriptive approaches employed. Also, these aspects often combine very specific details with more general guidelines or indicate intents and rationales without clarifying their implementation. In this paper we provide a method to describe immersive learning cases that is structured to enable comparisons, yet flexible enough to allow researchers and practitioners to decide which aspects to include. This method leverages a taxonomy that classifies educational aspects at three levels (uses, practices, and strategies) and then utilizes two frameworks, the Immersive Learning Brain and the Immersion Cube, to enable a structured description and interpretation of immersive learning cases. The method is then demonstrated on a published immersive learning case on training for wind turbine maintenance using virtual reality. Applying the method results in a structured artifact, the Immersive Learning Case Sheet, that tags the case with its proximal uses, practices, and strategies, and refines the free text case description to ensure that matching details are included. This contribution is thus a case description method in support of future comparative research of immersive learning cases. We then discuss how the resulting description and interpretation can be leveraged to change immersion learning cases, by enriching them (considering low-effort changes or additions) or innovating (exploring more challenging avenues of transformation). The method holds significant promise to support better-grounded research in immersive learning.
Candidate young stellar objects in the S-cluster: Kinematic analysis of a sub...Sérgio Sacani
Context. The observation of several L-band emission sources in the S cluster has led to a rich discussion of their nature. However, a definitive answer to the classification of the dusty objects requires an explanation for the detection of compact Doppler-shifted Brγ emission. The ionized hydrogen in combination with the observation of mid-infrared L-band continuum emission suggests that most of these sources are embedded in a dusty envelope. These embedded sources are part of the S-cluster, and their relationship to the S-stars is still under debate. To date, the question of the origin of these two populations has been vague, although all explanations favor migration processes for the individual cluster members. Aims. This work revisits the S-cluster and its dusty members orbiting the supermassive black hole SgrA* on bound Keplerian orbits from a kinematic perspective. The aim is to explore the Keplerian parameters for patterns that might imply a nonrandom distribution of the sample. Additionally, various analytical aspects are considered to address the nature of the dusty sources. Methods. Based on the photometric analysis, we estimated the individual H−K and K−L colors for the source sample and compared the results to known cluster members. The classification revealed a noticeable contrast between the S-stars and the dusty sources. To fit the flux-density distribution, we utilized the radiative transfer code HYPERION and implemented a young stellar object Class I model. We obtained the position angle from the Keplerian fit results; additionally, we analyzed the distribution of the inclinations and the longitudes of the ascending node. Results. The colors of the dusty sources suggest a stellar nature consistent with the spectral energy distribution in the near and midinfrared domains. Furthermore, the evaporation timescales of dusty and gaseous clumps in the vicinity of SgrA* are much shorter ( 2yr) than the epochs covered by the observations (≈15yr). In addition to the strong evidence for the stellar classification of the D-sources, we also find a clear disk-like pattern following the arrangements of S-stars proposed in the literature. Furthermore, we find a global intrinsic inclination for all dusty sources of 60 ± 20◦, implying a common formation process. Conclusions. The pattern of the dusty sources manifested in the distribution of the position angles, inclinations, and longitudes of the ascending node strongly suggests two different scenarios: the main-sequence stars and the dusty stellar S-cluster sources share a common formation history or migrated with a similar formation channel in the vicinity of SgrA*. Alternatively, the gravitational influence of SgrA* in combination with a massive perturber, such as a putative intermediate mass black hole in the IRS 13 cluster, forces the dusty objects and S-stars to follow a particular orbital arrangement. Key words. stars: black holes– stars: formation– Galaxy: center– galaxies: star formation
Authoring a personal GPT for your research and practice: How we created the Q...Leonel Morgado
Thematic analysis in qualitative research is a time-consuming and systematic task, typically done using teams. Team members must ground their activities on common understandings of the major concepts underlying the thematic analysis, and define criteria for its development. However, conceptual misunderstandings, equivocations, and lack of adherence to criteria are challenges to the quality and speed of this process. Given the distributed and uncertain nature of this process, we wondered if the tasks in thematic analysis could be supported by readily available artificial intelligence chatbots. Our early efforts point to potential benefits: not just saving time in the coding process but better adherence to criteria and grounding, by increasing triangulation between humans and artificial intelligence. This tutorial will provide a description and demonstration of the process we followed, as two academic researchers, to develop a custom ChatGPT to assist with qualitative coding in the thematic data analysis process of immersive learning accounts in a survey of the academic literature: QUAL-E Immersive Learning Thematic Analysis Helper. In the hands-on time, participants will try out QUAL-E and develop their ideas for their own qualitative coding ChatGPT. Participants that have the paid ChatGPT Plus subscription can create a draft of their assistants. The organizers will provide course materials and slide deck that participants will be able to utilize to continue development of their custom GPT. The paid subscription to ChatGPT Plus is not required to participate in this workshop, just for trying out personal GPTs during it.
The cost of acquiring information by natural selectionCarl Bergstrom
This is a short talk that I gave at the Banff International Research Station workshop on Modeling and Theory in Population Biology. The idea is to try to understand how the burden of natural selection relates to the amount of information that selection puts into the genome.
It's based on the first part of this research paper:
The cost of information acquisition by natural selection
Ryan Seamus McGee, Olivia Kosterlitz, Artem Kaznatcheev, Benjamin Kerr, Carl T. Bergstrom
bioRxiv 2022.07.02.498577; doi: https://doi.org/10.1101/2022.07.02.498577
Anti-Universe And Emergent Gravity and the Dark UniverseSérgio Sacani
Recent theoretical progress indicates that spacetime and gravity emerge together from the entanglement structure of an underlying microscopic theory. These ideas are best understood in Anti-de Sitter space, where they rely on the area law for entanglement entropy. The extension to de Sitter space requires taking into account the entropy and temperature associated with the cosmological horizon. Using insights from string theory, black hole physics and quantum information theory we argue that the positive dark energy leads to a thermal volume law contribution to the entropy that overtakes the area law precisely at the cosmological horizon. Due to the competition between area and volume law entanglement the microscopic de Sitter states do not thermalise at sub-Hubble scales: they exhibit memory effects in the form of an entropy displacement caused by matter. The emergent laws of gravity contain an additional ‘dark’ gravitational force describing the ‘elastic’ response due to the entropy displacement. We derive an estimate of the strength of this extra force in terms of the baryonic mass, Newton’s constant and the Hubble acceleration scale a0 = cH0, and provide evidence for the fact that this additional ‘dark gravity force’ explains the observed phenomena in galaxies and clusters currently attributed to dark matter.
CLASS 12th CHEMISTRY SOLID STATE ppt (Animated)eitps1506
Description:
Dive into the fascinating realm of solid-state physics with our meticulously crafted online PowerPoint presentation. This immersive educational resource offers a comprehensive exploration of the fundamental concepts, theories, and applications within the realm of solid-state physics.
From crystalline structures to semiconductor devices, this presentation delves into the intricate principles governing the behavior of solids, providing clear explanations and illustrative examples to enhance understanding. Whether you're a student delving into the subject for the first time or a seasoned researcher seeking to deepen your knowledge, our presentation offers valuable insights and in-depth analyses to cater to various levels of expertise.
Key topics covered include:
Crystal Structures: Unravel the mysteries of crystalline arrangements and their significance in determining material properties.
Band Theory: Explore the electronic band structure of solids and understand how it influences their conductive properties.
Semiconductor Physics: Delve into the behavior of semiconductors, including doping, carrier transport, and device applications.
Magnetic Properties: Investigate the magnetic behavior of solids, including ferromagnetism, antiferromagnetism, and ferrimagnetism.
Optical Properties: Examine the interaction of light with solids, including absorption, reflection, and transmission phenomena.
With visually engaging slides, informative content, and interactive elements, our online PowerPoint presentation serves as a valuable resource for students, educators, and enthusiasts alike, facilitating a deeper understanding of the captivating world of solid-state physics. Explore the intricacies of solid-state materials and unlock the secrets behind their remarkable properties with our comprehensive presentation.
(June 12, 2024) Webinar: Development of PET theranostics targeting the molecu...Scintica Instrumentation
Targeting Hsp90 and its pathogen Orthologs with Tethered Inhibitors as a Diagnostic and Therapeutic Strategy for cancer and infectious diseases with Dr. Timothy Haystead.
Discovery of An Apparent Red, High-Velocity Type Ia Supernova at 𝐳 = 2.9 wi...Sérgio Sacani
We present the JWST discovery of SN 2023adsy, a transient object located in a host galaxy JADES-GS
+
53.13485
−
27.82088
with a host spectroscopic redshift of
2.903
±
0.007
. The transient was identified in deep James Webb Space Telescope (JWST)/NIRCam imaging from the JWST Advanced Deep Extragalactic Survey (JADES) program. Photometric and spectroscopic followup with NIRCam and NIRSpec, respectively, confirm the redshift and yield UV-NIR light-curve, NIR color, and spectroscopic information all consistent with a Type Ia classification. Despite its classification as a likely SN Ia, SN 2023adsy is both fairly red (
�
(
�
−
�
)
∼
0.9
) despite a host galaxy with low-extinction and has a high Ca II velocity (
19
,
000
±
2
,
000
km/s) compared to the general population of SNe Ia. While these characteristics are consistent with some Ca-rich SNe Ia, particularly SN 2016hnk, SN 2023adsy is intrinsically brighter than the low-
�
Ca-rich population. Although such an object is too red for any low-
�
cosmological sample, we apply a fiducial standardization approach to SN 2023adsy and find that the SN 2023adsy luminosity distance measurement is in excellent agreement (
≲
1
�
) with
Λ
CDM. Therefore unlike low-
�
Ca-rich SNe Ia, SN 2023adsy is standardizable and gives no indication that SN Ia standardized luminosities change significantly with redshift. A larger sample of distant SNe Ia is required to determine if SN Ia population characteristics at high-
�
truly diverge from their low-
�
counterparts, and to confirm that standardized luminosities nevertheless remain constant with redshift.
Discovery of An Apparent Red, High-Velocity Type Ia Supernova at 𝐳 = 2.9 wi...
A biotechnology dream nitrogen fixing cereal crops by Deepak Sharma
1. A biotechnology dream: nitrogen fixing
cereal crops
Speaker: Sharma Deepak D.
Reg. no.: 1010119041
Major guide: Dr. V. P. Patel
Minor guide: Dr. V. B. Parekh
Course no: MBB-692
Date: 16/07/2021
Time: 3:00- 4:00 pm
2. Introduction
Biological Nitrogen Fixation
Mechanism of Symbiosis
Mechanism of Nitrogen fixation
Strategies to Transfer Symbiotic Nitrogen Fixation to Non-leguminous Crops
Case studies
Conclusion
2
Outline of the seminar
2
3. Introduction
The agriculture sector contributes 64% of world’s economic production.
Provides employment to 53% of the population of India (April, 2017).
Cereals like rice, wheat, maize, sorghum and millets are the crops with total
annual yields of 2000 million tons whereas two-third population consumes
only wheat worldwide
The world population today is 7.7 billion of whom 3.84 billion
people are fed by synthetic fertilizers.
World population is estimated to rise to 8.6 billion by 2030 and we
would need 185 mha more harvested area to feed this population.
The global nitrogen demand has been increasing by 1.3% p.a. and is
estimated to reach 132 mt by 2030 (101 mt in 2010).
3
4. • It represents about 2 % of the total plant
dry matter that enters the food chain.
• Nitrogen is an essential macronutrient
and is a component of proteins, nucleic
acids, and the energy currency of cells,
ATP.
• It is a component of the photosynthetic
pigment chlorophyll.
• It is a component of other important
alkaloids, hormones, etc.
Fig. 1. Nitrogen containing compounds.
(Buchanan et al., 2015)
4
Role of nitrogen in plants
5. Fig. 2. Relative contribution of the main crop categories to total N fertilizer
consumption (2018) in the main fertilizer markets. (Heffer, 2019)
Year 2018 2019 2020
Nitrogen (N) 115 376 117 116 118 763
Phosphate (P2O5 44 120 45 013 45 858
Potash (K2O) 34 894 35 978 37 042
Total (N+ P2O5 +K2O) 194 390 198 107 201 663
Table 1. World demand for fertilizer nutrient use, 2018-20 (000’ tonnes) (FAO, 2020)
5
6. Chemical Fertilizers
Urea (46%)
Ammonium Nitrate (33-
34%)
Di-ammonium Phosphate
(18%)
Organic Fertilizers
Bulky Organic Manure
1) FYM – 0.5%
2) Sheep and Goat Manure – 3%
3) Poultry Manure – 3.03%
Concentrated Organic Manure
1) Oilcakes – Mahua cake – 2.5%
Safflower cake – 7.9% N
2) Animal based Manures (meals)
Steamed bone meal – 1-2%
Horn and Hoof meal – 13%
Natural Sources
Fixation by lightening
Biological nitrogen
fixation
Sources of nitrogen in agriculture
6
7. Fig. 3. Contribution of various sources to the
fixed nitrogen in soil
(Buchanan et al. 2015) 7
9. Biological Nitrogen Fixation is the fixation of elemental dinitrogen (N2), from
the atmosphere, by soil micro organisms through a reductive process into
ammonia.
Fig. 4. Primary nitrogen fixing genera
(Rogers and Oldroyd, 2014)
Biological Nitrogen Fixation
9
10. (Mus et al., 2016)
Fig. 5. Schematic representation of the different associations
between diazotrophs and plant hosts. 10
11. Fig. 6. Plant-microbe interaction in the soil
Flavone Chalcone Isoflavone
Fig. 7. Diverse structures of flavonoids
(Buchanan et al.,2015) 11
Mechanism of symbiosis
Rhizobial Symbiosis
12. Fig. 8. Perception of Nod Factors by plant membrane
receptors
Contd.
Cell membrane
Nuclear
membrane
(Rogers and Oldroyd, 2014) 12
• The rhizobium–legume symbiosis is
initiated by flavonoids released from the
plant roots that stimulate rhizobia to
produce the signalling molecule Nod
factor.
• The perception of Nod factor drives
developmental responses in the compatible
plant host that prepares the plant to
accommodate the endosymbiotic partner.
• Perception of Nod factor initiates two
coordinated genetic programmes: the
initiation of cell divisions in the root cortex
and the development of an infection thread
(or IT) in the root epidermis.
13. Fig. 9. Regulation by CCaMk
Contd.
(Buchanan et al.,2015) 13
• Nodulation signalling downstream of calcium spiking.
• Calcium spiking is perceived by the binding of calcium and calmodulin (CaM) to
CCaMK.
15. Fig. 11. Events that lead to nitrogen‐fixing cells.
Cont.
(Buchanan et al., 2015) 15
16. Fig. 12. Structures of MoFe and Fe proteins of nitrogenase, and electron flow
through the two enzymes. (Buchanan et al., 2015)
nifH
nifDK
16
Mechanism of Nitrogen Fixation
17. A) Components I and II are dissociated; II is
ready for reduction.
B) ATP binds to component II, which receives
electrons from an electron donor
(ferredoxin or flavodoxin); binding of ATP
induces an allosteric conformational change
which allows association of the two
proteins. Electrons flow from the [4Fe-4S]
cluster on II to the P cluster on I.
C) Electrons are further shuttled to the iron-
molybdenum cofactor (FeMoco), and ATP
is hydrolised to ADP. This step is repeated
several times before a molecule of N2 can
bind to FeMoco.
D) The protein complex dissociates, and
nitrogenase reduces dinitrogen to ammonia
and dihydrogen..
Fig. 13. A general catalytic mechanism scheme for nitrogenase
17
18. Effect of Oxygen
18
Fig. 15. Gene regulation based on free
oxygen concentration
Fig. 14. Mechanisms to maintain low free
oxygen.
2) Spatial or temporal separation of
photosynthesis and nitrogen fixation.
1)
(Buchanan et al. 2015) 18
21. Fig. 18. Association of diazotrophs with plants as a potential gateway to sustainable
agriculture: strategies, tools, and challenges for engineering symbiotic nitrogen fixation.
Strategies to transfer SNF to Non-leguminous crops
Identify the gens in CSP
and use them to establish
RNS
Inoculate non-legumes
with endophytic
diazotrophs
Engineer nitrogen fixing
plants
Use endophytic bacteria as
a chassis for nitrogen
fixation
Create a biased rhizosphere
to encourage transkingdom
signalling
(Mus et al., 2016)
21
22. Table. 2. Major genomic loci detected for BNF in different legume species
22
23. Table 3. Functions of NOD, NIF, and FIX genes
Gene Function References
nod gene
nod M Nod factor synthesis Merrick (1992)
nod L Determine host range Gottfert (1993)
nod E Determine host range Gottfert (1993)
nod F Encodes a specific acyl carrier protein used to acylatethe nod factor specified by nod A Merrick (1992)
nod D Encodes a regulatory protein , Nod D that controlstranscription of other nod genes Merrick (1992)
nod A Direct synthesis of nod factor backbone Gottfert (1993)
nod B Direct synthesis of nod factor backbone Gottfert (1993)
nod C Direct synthesis of nod factor backbone Gottfert (1993)
nod I Membrane proteins that help in exporting nod factors Kondorosi et al. (1991)
nod J Membrane proteins that help in exporting nod factors Kondorosi et al. (1991)
fix gene
fix LJ Oxygen-responsive two-component regulatory system involved in positive control of
fixK and nifA
David et al. (1988)
fix K Positive regulator of fixNOQP, nifA; negative regulatorof nifA and fixK Batut et al. (1989)
fix NOQP Microaerobically induced, membrane-bound cyto-chrome oxidase Boistard et al. (1991)
fix GHIS Redox process-coupled cation pump Kahn et al. (1989)
fix ABCX Unknown function; required for nitrogenase activity; FixX shows similarity to
ferredoxins
Earl et al. (1987)
fix R Unknown function; not essential for nitrogen fixation Thony et al. (1987) 23
24. nif gene
nifD
α subunit of dinitrogenase. Forms an α2 ß2 tetramer with ß subunit interface. FeMo-co, the site substrate
reduction, is present buried within the α subunit of dinitrogenase
Dean and Jacobson (1992)
nifK ß subunits of dinitrogenase. ß clusters are present at ßsubunit interface
nifH
Fe protein subunit of dinitrogenase reductase. Obligate electron donor to dinitrogenase during
dinitrogenaseturnover. Also is required for FeMo-co biosynthesis
nifN Required for FeMo cofactor biosynthesis Allen et al. (1994)
nifV Encodes a homocitrate synthase. Homocitrate is anorganic component of FeMo cofactor Hoover et al. (1987)
nifB
Required for FeMo cofactor biosynthesis. Metabolicproduct. NifB-co is the specific Fe and S donor to
FeMo-co
Shahet al. (1999)
nifQ Incorporation of Mo into FeMo cofactor. Proposed tofunction in early MoO 2 processing Imperial et al. (1984)
nifE Forms ɑ2ß2 tetramer with nifN. Required for FeMocofactor biosynthesis Allen et al. (1994)
nifX Not essential for nitrogen fixation; required for FeMocofactor biosynthesis Shah et al. (1999)
nifU Involved in mobilization of Fe-S cluster synthesis andrepair Yuvaniyama et al. (2000)
nifS Involved in mobilization of Fe-S cluster synthesis andrepair Zheng et al. (1993)
nifY Associates with MoFe protein and dissociates uponFeMo cofactor insertion Homer et al. (1993)
nifM
Required for the maturation of nifH and Fe protein maturation. Putative peptidyl-prolyl cis/trans
isomerase
Dean and Jacobson (1992)
nifW Involved in stability of dinitrogenase. Proposed to pro-tect dinitrogenase from O inactivation Kim and Burgess (1996)
nifF Flavodoxin required for electron transfer to the Feprotein, Physiologic electron donor to nifH Thorneley et al. (1992)
nifJ
Pyruvate flavodoxin (ferredoxin) oxidoreductase involved in electron transport to nitrogenase; couples
pyruvate oxidation to reduction of the nifF product
Shah et al. (1988)
nifA Positive regulator of nif transcription Dixon (1998)
nifL Negative regulatory protein Dixon (1998) 24
Cont.
26. (Liu et al., 2018)
(Washington University, USA)
Case study I
Objective
• Engineer nitrogenase activity in the nondiazotrophic oxygenic photosynthetic cyanobacterium Synechocystis
sp. PCC 6803 through the transfer of 35 nitrogen fixation (nif) genes from the diazotrophic cyanobacterium
Cyanothece sp. ATCC 51142.
26
27. Materials and methods
Microorganisms, culture conditions, and media
• All cyanobacterial strains, including Cyanothece 51142,
Synechocystis 6803, and engineered strains were
cultured BG11 medium
• Yeast and E. coli strains yeast extract-peptone-dextrose
plus adenine (YPAD) medium and LB medium resp.
Construction of recombinant plasmids and
engineered strains
• Plasmids constructed by DNA assembler and Gibson
assembly.
• The sequences of all of the plasmids constructed in this
study were verified (Genewiz, NJ).
• pSyNif-1 and pSyNif-2 were introduced into the WT
strain through triparental conjugation and other
recombinant plasmids by natural transformation.
RT-PCR and q-PCR.
• RNA samples from culture (BG110) used for RT-PCR. cDNA
generated after reverse transcription was used as the template for PCR
to validate the transcription of genes.
• q-PCR was performed on RNA samples extracted from culture grown
in BG110 medium under light/dark conditions.
Measurement of nitrogen fixation activity
• Nitrogen fixation activity was measured by an Acetylene Reduction Assay
In vivo 15N2 incorporation assay
Isotope ratios were measured by elemental analyzer-
isotope ratio mass spectrometry and values are indicated as
Δ15N %, where the number represents a linear transform of
the 15N/14N isotope ratios.
Western blot analysis.
• PCR amplification of nifH gene and nifD gene from the genomic
DNA of Cyanothece 51142.
• The protein purified from E. coli BL21 (DE3) transformed with
plasmids pET28a-nifH and pET28a-nifD used as positive control.
• Proein extracted from Cyanobacterial cells cultured in N-free medium
and Immunodetection was performed using Western blotting Luminol
reagent.
27
28. Introduction of nif genes into Synechocystis 6803
(A) Phylogeny of cyanobacteria. (B) Genetic organization of the nif cluster and (C) the role of each gene product in Cyanothece 51142: Three structural
proteins (nifHDK; green), necessary cofactors (blue), accessory proteins (orange), ferredoxins (purple), and hypothetical proteins (brown). (D) Plasmid
pSyNif-1 containing the entire nif cluster. The backbone (gray) is from broad host plasmid pRSF1010, which can replicate in Synechocystis 6803. The yeast
helper fragment (black) contains CEN6 and ARS as an ori and ura3 as a selection marker. (E) Transcription of all 35 genes in engineered Synechocystis 6803.
Fig. 19. Introduction of nif genes into Synechocystis 6803.
Results
28
29. Scheme showing the top-down method to determine the
minimal nif cluster.
Fig. 20. The minimal nif cluster required for nitrogen fixation activity in Synechocystis 6803.
Nitrogen fixation activity in engineered strains.
The minimal required gene cluster for nitrogen fixation activity
29
30. • To increase the RNA expression levels of the
nitrogenase-related genes,
• Three small endogenous plasmids in Synechocystis
6803: pCA2.4, pCB2.4, and pCC5.2 maintained
higher transcriptional levels than those in a
pRSF1010-based plasmid, because of the higher
copy numbers of these three plasmids within
Synechocystis
• By replacing RSF1010 backbone of plasmid pSyNif-
2 by these endogenous episomes and then transferred
the plasmids to Synechocystis 6803,
• Generating three strains, TSyNif-9, TSyNif-10, and
TSyNif-11, with the chassis of pCA2.4, pCB2.4, and
pCC5.2, respectively.
• Genes nifH, nifD, and nifK in strain TSyNif-9
showed transcription levels that were several fold
higher than in TSyNif-2. In addition, nitrogen
fixation activity was increased by another 2- to 3-
fold,
Fig. 21. Enhancement of transcription levels of nif genes leads to higher
nitrogen fixation activity.
Improvement of nitrogen fixation activity.
30
31. Fig. 22. Expression of uptake hydrogenase improves O2 tolerance
of nitrogenase.
Improvement of O2 tolerance by introduction of
hydrogenase uptake
• In order to test the oxygen sensitivity of nitrogen fixation
activity in TSyNif-2, a measured amount of oxygen was
added to the headspace of cultures grown in BG110 media
to generate micro-oxic conditions of 0.5% and 1.0% of O2
in the sealed testing bottles. The activity dropped more than
10-fold and 60-fold resp.
• To enhance O2 tolerance under the same conditions, genes
coding for the uptake hydrogenase enzyme from
Cyanothece 51142 were introduced into the chromosome of
the TSyNif-2 strain.
• The structural genes for this hydrogenase, hupS and hupL,
present together in a single operon in Cyanothece 51142,
were transformed into TSyNif-2, generating strain TSyNif-
12
• HupW is required for the maturation of HupL protein.Thus,
the hupSLW genes organized in two operons were
transformed into TSynif-2 to generateTSyNif-13.
• The expression of hup genes in TSyNif-12 and TSyNif-13
was assessed by RT-PCR
31
32. (Geddes et al., 2019)
(University of Oxford, UK)
Case study II
Objective:
• To engineer barley plants that exude scyllo-inosamine (SIA) into the rhizosphere.
• To establish synthetic SIA-mediated transkingdom signalling between transgenic barley plants and R.
leguminosarum.
32
33. Methods
Bacterial strains and growth media
• Escherichia coli strains were grown in
Luria–Bertani (LB) medium
• Rhizobium leguminosarum and
Sinorhizobium meliloti strains were
grown in tryptone yeast (TY) medium
or universal minimal salts (UMS)
Plant materials and sterilization
• Medicago sativa, Pisium sativum,
Medicago truncatula ecotype
Jester and barley seeds were
sterilized with standard protocol.
Bacterial genetic manipulations and plasmid construction.
• All plasmids were verified by restriction digest and DNA
sequencing. Plasmids were transferred by conjugation into rhizobia
by tri-parental mating with pRK2013.
Generation of constructs for plant engineering.
• For transient expression, R. leguminosarum idhA and S. meliloti L5-
30 mosB were expressed under the control of CaMV 35s promoter
and Lotus japonicus Ubiquitin1 (LjUBI1) promoter, respectively.
• S. meliloti mosDEF and M. crystallinum IMT were expressed
transiently under the control of 35s and LjUBI1 promoters,
respectively.
• In barley transgenic lines, idhA and mosB were expressed under the
control of Zea mays ubiquitin1 promoter and Oryza sativa ubiquitin1
promoter, respectively.
• Constructs for plant engineering were assembled using golden gate
cloning. Recombinant clones were verified by PCR and DNA
sequence analysis.
33
34. Metabolite extraction and GC-MS analysis of root nodules.
• Ten germinated M. sativa or two germinated P. sativum seedlings
per pot were inoculated with 1 × 107 colony-forming units (CFUs)
of rhizobia in distilled water 3 days after planting.
• Plants were grown for 6 to 8 weeks before harvesting pink N2-
fixing nodules by hand.
• For metabolite extraction, 750 μl of CHCl3 and 1400 μl of dH2O
was added, samples were vortexed and then centrifuged for 15 min
at a relative centrifugal force (RCF) of 2200.
• GC-MS analysis was performed using the LJS_TMSI protocol at
the University of Oxford Department of Plant Sciences. GC-MS
was performed with the LJS Golm Stardard protocol.
Protein purification
• Purification of HIS-tagged proteins
from the lysate was performed
using a His-Spin Protein Miniprep
Kit (Zymo Research).
• Purity of purified proteins was
assessed by sodium dodecyl sulfate-
polyacrylamide gel electrophoresis
analysis and proteins were
quantified using a Qubit 2.0
Fluorometer (Thermo Fisher)
according to the manufacturer’s
instructions.
Transient expression in Nicotiana benthamiana.
• Recombinant plasmids were mobilised into A. tumefaciens GV3101:pMP90 by electroporation.
• A transformed single colony was grown in LB. A. tumefaciens strains were mixed with an equal volume of a P19
suppressor strain and infiltrated into the underside of N. benthamiana leaves using a needleless 1 ml syringe.
• Leaf discs were harvested 3 days after infiltration, frozen in liquid nitrogen and the extracted metabolites were analysed
by GC-MS.
34
35. Hairy-root transformation in Medicago truncatula.
• A. rhizogenes AR1193 was transformed with
recombinant constructs by electroporation.
• Germinated seedlings of M. truncatula cultivar
Jester were transformed with A. rhizogenes
AR1193 using standard protocols.
Stable transformation of Hordeum vulgare.
• Recombinant binary vectors were transferred by
electroporation into A. tumefaciens AGL1 strain.
GC-MS analysis of engineered plant tissue.
• GC-MS analysis was performed using the
Eng_Plant protocol
• MassHunter qualitative analysis software
(Agilent) was used to determine the peak area of
rhizopine in EIC (m/z 245) chromatograms of the
standards and plant samples.
Bacterial luciferase biosensor screening assays
• Rhizopine lux biosensor was inoculated on the
engineered roots and plants were imaged each
day post-inoculation.
• Bioluminescence images were analysed for
quantification using imaging software IndiGO
(Berthold Technologies) and data were
expressed as the ratio of luminescence to
surface (cps mm−2).
Analysis of growth and bioreporter expression
in culture.
• Analysis of expression of bioreporters in free-
living culture was performed with either a
FLUOstar OMEGA (Lite) or CLARIOstar
plate reader (BMG Labtech).
• Data are expressed as relative luminescence
units calculated from total luminescence per
well/ culture density measured by OD595.
35
36. Results
Rhizopine exudation during rhizobial
symbiosis
a) Chemical structures of rhizopines and
related cyclitols.
b) NightOwl images of bioluminescence
of R. leguminosarum Rlv3841/pOPS0046
rhizopine lux biosensor on the surface of
M. sativa roots nodulated by S. meliloti
L5-30 wild-type (rhizopine+) and S.
meliloti L5-30 mosB:pK19 (rhizopine –,
Fig. 2a).
c) Specificity of induction of rhizopine
lux biosensor with rhizopines compared to
other chemically similar plant polyols
d) Induction curves showing the dynamic
range and sensitivity of the rhizopine lux
biosensor with chemically synthesised
SIA 1. Fig. 23. Detection of rhizopine exudation with a rhizopine biosensor 36
37. Fig. 24. Discovery of a natural and synthetic pathway for rhizopine synthesis.
37
a) Metabolites from nodules formed
by wild-type S. meliloti L5-30
compared to mosB:pK19 and
mosE:pK19 mutants.
b) Wild-type S. meliloti Rm1021
maintaining an empty vector
(EV) and expressing mosDEF.
c) GC-MS chromatograms (TIC) of
MosB in vitro assay in the
absence of protein or with MosB
added.
d) Proposed natural pathway of 3-O-
MSI biosynthesis in S. meliloti
L5-30
e) Linked in vitro assay of inositol
dehydrogenase IolG and MosB
f) Extracts prepared from tobacco
leaves agroinfiltrated with either
empty vector (control) or IdhA or
MosB or IdhA-MosB together
g) Proposed synthetic pathway of
SIA 1 synthesis
38. • a, b GC-MS TICs of extracts prepared
from M. truncatula transgenic roots (a)
and transgenic T0 barley seedlings (b)
transformed with empty vector (control)
or IdhA-MosB together.
• Highlighted peaks indicate scyllo-
inosamine 1 (orange), scyllo-inosose 7
(green) and myo-inositol 3 (dark blue).
• c, d NightOwl images showing
bioluminescence of Rlv3841/pOPS0046
rhizopine lux biosensor on the surface of
M. truncatula transgenic roots (c) and T0
barley seedlings (d) transformed with
empty vector (control) or IdhA-MosB
together (engineered).
Fig. 25. Rhizopine biosynthesis and signalling in M. truncatula and barley roots 38
39. a) Confocal fluorescence microscopy images showing
green fluorescent protein (GFP) fluorescence of
Rlv3841::mTn7-mCherry/pOPS0761 biosensor [with
constitutive mCherry fluorescence] on the surface of T1
barley seedlings transformed with empty vector
(control) or IdhA-MosB together (engineered).
Z-stack projections of green fluorescence channel (left,
GFP), red fluorescence channel (middle, mCherry) and
all channels merged (right, bright-field) are shown
(scale bars, 100 µm).
b) Three-dimensional images of GFP/mCherry mean
intensity ratios in Rlv3841::mTn7-mCherry/pOPS0761
biosensor on the surface of T1 barley seedlings
transformed with empty vector (left, control) or IdhA-
MosB together (middle, engineered).
Cool colours (purple) indicate low GFP/mCherry
intensity ratio, and warm colours (red) indicate a high
GFP/mCherry intensity ratio.
Fig. 26. Fluorescent microscopy of rhizopine-mediated
transkingdom signalling
39
40. (Bloch et al., 2020)
(Berkeley, USA)
Case study III
Objective
• Identification and isolation of diazotrophs that closely associate with key crops to improve nitrogen
contributions by BNF
• Gene editing to disrupt regulatory networks linking nitrogen sensing, fixation, and assimilation
40
41. Materials and methods
Isolation of Kosakonia sacchari PBC6.1
• Isolated bacterial Colonies from corn seedling
that emerged were tested for the presence of the
nifH gene by colony PCR
Fluorescence microscopy
• PBC6.1 was transformed with plasmid PB114-
RFP, a plasmid containing the pSC101 origin of
replication, chloramphenicol resistance cassette,
and a gene encoding red fluorescent protein (RFP)
under the control of a strong constitutive promoter.
• Transformed cell suspension was inoculated
directly on the seed at the time of seeding.
• Root sections were imaged on a 6D Widefield
Nike TI inverted fluorescence microscope
Initial field trial with isolated diazotrophs
• Corn seed was coated with a culture suspension of
6 strains selected from 49 strains isolated
• Root samples were processed and assayed for
colonization of the inoculant diazotrophs
Extraction of the root-associated microbiome
• Genomic DNA extraction was performed with the ZR-96 Quick-
gDNA kit, and RNA extraction was performed using the RNeasy kit.
Root colonization assay
• Quantification of root colonization was done by using quantitative
PCR. For each experiment, the colonization numbers were compared
with UTC seedling
Acetylene reduction assay (ARA)
• A modified version of the ARA (Temme et al., 2012) was used to
measure nitrogenase activity in pure culture conditions.
Ammonium excretion assay
• Supernatant from the reactor broth was assayed for free ammonium
using the Megazyme Ammonia Assay kit normalized to biomass at
each time point.
41
42. Gene editing
• The genome modifications described in Fig. 2 were
generated using genome editing methods described in
a recently published patent application on our guided
microbial remodeling platform (Bloch et al., 2019b).
• Genome-edited strains were cured of all plasmids used
to carry out genome editing by repeated subculturing
followed by sequence verification of the desired edits.
Greenhouse assays to measure nifH transcription
in PBC6.29, PBC6.99, PBC6.38, and PBC6.94
• Plants grown in nitrogen free background. Each seed was
inoculated with either sterile phosphate-buffered saline
(controls) or an equal volume of microbial suspension
using cells diluted to a set OD.
• At 2 or 4 weeks after planting, plants were harvested, and
nucleic acids were extracted from root tissue.
Colonization of each strain was measured.
• For microbial RNA quantification, microbial RNA was
extracted using the RNeasy Kit (Qiagen) The extracted
RNA was used for NanoString analysis of nifA, nifH, and
rpoB microbial transcript on an nCounter Sprint.
• Field trials to measure colonization and nifH
transcription were carried out at California, Puerto Rico
and Illinois
Re-isolation of edited strains from field corn
root samples
• Isolates were screened using PCR primers specific to
the mutations, and those with the correct band sizes
were further verified by Sanger sequencing of the 16S
rRNA region.
• Between two and 20 clones of each re-isolated strain
were then purified and assessed for ARA activity
relative to the original inoculant strain
42
43. Results
Fig. 27. Kosakonia sacchari PBC6.1 contains well-
characterized nitrogen regulatory pathways.
• PBC6.1 has a genome of at least 5.4 Mbp, and a nif gene cluster and a
nitrogen metabolic regulatory network.
• The nifLA operon directly regulates the rest of the nif cluster through
transcriptional activation by NifA and nitrogen- and oxygen-dependent
repression of NifA by NifL.
• Glutamine synthetase (GS) is responsible for rapid assimilation of newly
fixed nitrogen in nitrogen limiting conditions.
• GlnE- adenylylation- attenuate or deadenylylation- restore activity
• The nifLA operon and GS (encoded by the glnA gene) are regulated by
the PII protein regulatory cascade,
• No nitrogen- GlnD modifies PII proteins, leads to the up-regulation of
nitrogen fixation and assimilation pathways.
• Exogenous nitrogen- GlnD removes the covalent modification from the
PII proteins, leading to the repression of nitrogen fixation and
assimilation genes
43
44. Fig. 28. Mutated sequences of the key genes of the nitrogen fixation and
assimilation regulatory network of PBC6.1.
Remodel the regulatory networks of PBC6.1
1) Disruption of nifL can abolish inhibition of NifA and improve
nif expression in the presence of both oxygen and exogenous
fixed nitrogen, expressing nifA under the control of a
nitrogen-independent promoter could decouple nitrogenase
biosynthesis from the PII protein regulatory cascade (Fig. 2a)
2) Truncation of the GlnE protein to delete its adenylyl-removing
(AR) domain would lead to constitutively adenylylated GS,
limiting ammonium assimilation by the microbe and leading
to ammonium build up and release (Fig. 2b)
3) Abolishing expression of AmtB, the transporter responsible
for uptake of ammonium, could lead to greater extracellular
ammonium by preventing reuptake of excreted ammonium
(Fig. 2c)
4) Deletion of the GlnD protein would lead to a constant nitrogen
sufficiency signal by eliminating the ability of the cells to
covalently modify PII proteins (Fig. 2d).
5) Strain with the nifH gene deleted to serve as a negative control
for nitrogenase expression (Fig. 2e).
44
45. Table 4. List of isolated and remodeled K. sacchari strains used in this work
Strain Genotype
PBC6.1 WT
PBC6.15 ΔnifL::Prm5
PBC6.29 ΔnifL::Prm5 ΔglnEAR1
PBC6.99 ΔnifL::Prm5 ΔglnEAR1 ΔglnD
PBC6.90 ΔnifL::Prm5 ΔglnEAR1 ΔnifH
PBC6.14 ΔnifL::Prm1
PBC6.37 ΔnifL::Prm1 ΔglnE AR2
PBC6.38 ΔnifL::Prm1 ΔglnEAR1
PBC6.93 ΔnifL::Prm1 ΔglnE AR2 ΔamtB
PBC6.94 ΔnifL::Prm1 ΔglnE AR1 ΔamtB
45
46. Fig. 29. Edited strains of PBC6.1 fix nitrogen independently of nitrogen status.
• To assess the sensitivity of
PBC6.1 to exogenous nitrogen
and to predict activity in a
fertilized field, nitrogenase
activity in pure culture was
measured with the classical ARA
in the presence and absence of
fixed nitrogen.
• The wild-type strain exhibits
repression of nitrogenase activity
as glutamine concentrations
increase, while remodeled strains
show nitrogenase activity in the
presence of 5 mM glutamine
(Gln) or NH4
+.
• PBC6.90 was not tested in 5 mM
NH4
+; PBC6.99 was not tested in
5 mM glutamine.
46
47. Fig. 30. Edited strains of PBC6.1 excrete fixed nitrogen into their
environment.
• ∆nifL::Prm5 mutation alone (PBC6.15) was
insufficient to confer an ammonium excretion
phenotype, the ∆nifL::Prm1 mutation alone
(PBC6.14) led to significant excretion of ammonium.
• The ∆glnEAR mutations led to an increase in
ammonium excretion when stacked with the
∆nifL::Prm mutations, supporting our hypothesis that
down-regulation of GS activity would lead to
ammonium excretion
• The ∆glnD mutation led to an additional increase in
ammonium excretion, probably by causing a decrease
in glnA expression.
• The ammonium excretion phenotype conferred by the
∆glnEAR and ∆glnD mutations came with a
corresponding decrease in growth rates, similar to
what was observed in the acetylene reduction assay.
• The ∆amtB mutations had no apparent effect on
ammonium excretion or growth rate when stacked
with the ∆nifL::Prm and ∆glnEAR mutations.
• These results suggest that the edited strains may be
able to fix nitrogen and transfer it to the crop in
fertilized field conditions.
47
48. Fig. 31. Greenhouse experiments demonstrate rhizosphere nifH transcription in fertilized corn.
• To determine whether the
edited microbes were able
to express nitrogenase in
the rhizosphere of fertilized
plants, inoculated corn
plants in greenhouse assays
with PBC6.1 and a subset
of edited strains to measure
colonization of the
cornrhizosphere and nifH
transcription therein.
48
• black bars represent samples
analyzed using primers
targeting the nifH–por2
intergenic region, and hatched
bars represent samples
analyzed using primers
targeting the ∆nifL::Prm5
genotype
49. • Root samples were collected between 2
and 5 weeks after planting for nucleic
acid extraction to verify the presence of
the strain and expression of the nifH gene
• Root samples were collected, the root-
associated microbiome was extracted,
and nifH transcription (a, c, e, g) and
colonization (b, d, f, h) were quantified.
• At all locations, remodeled strains show
increased normalized nifH transcript
levels and similar colonization when
compared with PBC6.1.
• In (b, d, f, and h), black bars represent
samples analyzed using primers targeting
the nifH–por2 intergenic region; shaded
and hatched bars represent samples
analyzed using primers targeting
the ∆nifL::Prm1 and ∆nifL::Prm5
genotypes, respectively.
Fig 32. Remodeled strains colonize corn roots and express nitrogenase in diverse
locations, soil types, and nitrogen levels. 49
50. Fig 33. Corn inoculated with PBC6.1 and its derivatives exhibited
an increase in grain yield above UTC in the Puerto Rico field trial.
Fig. 34. Nitrogenase activity is observed in clones re-
isolated from field grown root samples.
50
51. • The creation of artificial symbioses or associations between diazotrophs and crops is a primary goal in
agriculture to reduce the demand for chemical nitrogen fertilizers
• Utilization of diazotrophs isolated from non-legumes in other non-leguminous crops has proved to be a
successful method to transfer nitrogen fixation
• Engineering nitrogen fixation activity in photosynthetic nondiazotrophic cyanobacteria paved the way to
resolve the problem of O2 sensitivity of nitrogenase enzyme.
• The minimal cluster for 24 nif genes for nitrogenase activity will provide a useful framework for refactoring
genes. But biosynthesis of fully functional nitrogenase is a complex process as genes with the same
designations in different species occasionally have alternative functions.
• The confirmation of the chemical structure of rhizopines, and their secretion into the rhizosphere, provides a
unique opportunity to use them as target molecules for engineering plant control of root bacteria (biased
rhizosphere)
• Rhizopine transkingdom signalling could control synthetic symbioses to deliver nitrogen to cereal crops.
• Through gene edited strains in which nitrogenase biosynthesis is decoupled from the regulatory networks that
sense and respond to cellular nitrogen status can fix and excrete significant quantities of nitrogen into their
environment at various levels of exogenous nitrogen is a first step toward developing strains that can replace
synthetic fertilizers in cereal crop production
Conclusion
51