Golden Rice is a humanitarian biotechnology project aimed at addressing vitamin A deficiency. It involves engineering rice to produce beta-carotene, a precursor to vitamin A, in the endosperm through the introduction of two transgenes - a phytoene synthase gene and a bacterial carotene isomerase gene. Extensive research over many years was needed to optimize beta-carotene levels, including testing different versions of phytoene synthase and selecting high expressing events. Human studies showed the beta-carotene in Golden Rice is bioavailable. Regulatory dossiers require both event independent and event dependent studies for approval prior to implementation.
This document discusses Agrobacterium tumefaciens, a soil bacterium that causes crown gall disease in plants. It transfers tumor-inducing (Ti) plasmid DNA into wounded plant cells, integrating it into the plant genome and causing tumors. The Ti plasmid contains the T-DNA region flanked by borders, which is transferred to plants. It also contains virulence genes and opine catabolism genes. The infection process involves signal induction, bacterial attachment, production of virulence proteins, T-DNA production and transfer into the plant cell nucleus where it integrates.
Golden rice is a variety of rice (Oryza sativa) produced through genetic engineering to biosynthesize beta-carotene, a precursor of vitamin A, in the edible parts of rice.It is intended to produce a fortified food to be grown and consumed in areas with a shortage of dietary vitamin A, a deficiency which each year is estimated to kill 670,000 children under the age of 5 and cause an additional 500,000 cases of irreversible childhood blindness. Rice is a staple food crop for over half of the world's population, providing 30–72% of the energy intake for people in Asian countries, and becoming an effective crop for targeting vitamin deficiencies.
This document provides information about abscisic acid (ABA), including its chemical structure, biosynthesis, roles in plants, and research findings. Some key points:
- ABA is a plant hormone involved in processes like seed dormancy, stomatal closure, and leaf senescence. It has a cis-trans isomer structure and exists primarily in the cis form.
- ABA is biosynthesized through direct and indirect pathways, with the indirect pathway being more common in plants. This pathway involves carotenoid precursors that are cleaved to form ABA.
- Research has examined ABA's role in regulating strawberry fruit development and ripening. Studies show ABA levels change over fruit growth stages and that
This document summarizes a presentation about a genetically engineered microorganism called a "super bug" developed to degrade hydrocarbons in petroleum waste. The super bug was constructed by Anand Chakrabarty et al. in 1979 through conjugative transfer of plasmids containing genes from Pseudomonas putida strains that degrade various hydrocarbons like camphor, octane, xylene, and naphthalenes. This created a strain with three plasmids allowing it to break down multiple pollutants. The super bug was selected and mass cultured, then used to treat oil spills by applying inoculated straw to spread the bacteria and degrade the oil over time.
This document summarizes information about Arabidopsis thaliana, a small flowering plant that is widely used as a model organism. It describes A. thaliana's physical characteristics and life cycle, genome, and use in understanding flower development. Specifically, it outlines the ABC model of flower development in which three classes of genes (A, B, and C) interact to specify the four types of floral organs - sepals, petals, stamens, and carpels. Mutations in these genes result in homeotic transformations where one organ develops in place of another.
Transposon mutagenesis & site directed mutagenesisAnuKiruthika
This document provides information on transposon mutagenesis and site-directed mutagenesis. It defines transposons as "jumping genes" that can move locations in the genome and explains that transposon mutagenesis uses transposons to cause mutations by interrupting genes. Site-directed mutagenesis is described as an in vitro technique to introduce specific mutations into DNA using methods like conventional PCR, nested PCR, or inverse PCR. The document outlines several applications of these mutagenesis methods such as identifying virulence genes or screening for desired mutations.
This document discusses Agrobacterium tumefaciens, a soil bacterium that causes crown gall disease in plants. It transfers tumor-inducing (Ti) plasmid DNA into wounded plant cells, integrating it into the plant genome and causing tumors. The Ti plasmid contains the T-DNA region flanked by borders, which is transferred to plants. It also contains virulence genes and opine catabolism genes. The infection process involves signal induction, bacterial attachment, production of virulence proteins, T-DNA production and transfer into the plant cell nucleus where it integrates.
Golden rice is a variety of rice (Oryza sativa) produced through genetic engineering to biosynthesize beta-carotene, a precursor of vitamin A, in the edible parts of rice.It is intended to produce a fortified food to be grown and consumed in areas with a shortage of dietary vitamin A, a deficiency which each year is estimated to kill 670,000 children under the age of 5 and cause an additional 500,000 cases of irreversible childhood blindness. Rice is a staple food crop for over half of the world's population, providing 30–72% of the energy intake for people in Asian countries, and becoming an effective crop for targeting vitamin deficiencies.
This document provides information about abscisic acid (ABA), including its chemical structure, biosynthesis, roles in plants, and research findings. Some key points:
- ABA is a plant hormone involved in processes like seed dormancy, stomatal closure, and leaf senescence. It has a cis-trans isomer structure and exists primarily in the cis form.
- ABA is biosynthesized through direct and indirect pathways, with the indirect pathway being more common in plants. This pathway involves carotenoid precursors that are cleaved to form ABA.
- Research has examined ABA's role in regulating strawberry fruit development and ripening. Studies show ABA levels change over fruit growth stages and that
This document summarizes a presentation about a genetically engineered microorganism called a "super bug" developed to degrade hydrocarbons in petroleum waste. The super bug was constructed by Anand Chakrabarty et al. in 1979 through conjugative transfer of plasmids containing genes from Pseudomonas putida strains that degrade various hydrocarbons like camphor, octane, xylene, and naphthalenes. This created a strain with three plasmids allowing it to break down multiple pollutants. The super bug was selected and mass cultured, then used to treat oil spills by applying inoculated straw to spread the bacteria and degrade the oil over time.
This document summarizes information about Arabidopsis thaliana, a small flowering plant that is widely used as a model organism. It describes A. thaliana's physical characteristics and life cycle, genome, and use in understanding flower development. Specifically, it outlines the ABC model of flower development in which three classes of genes (A, B, and C) interact to specify the four types of floral organs - sepals, petals, stamens, and carpels. Mutations in these genes result in homeotic transformations where one organ develops in place of another.
Transposon mutagenesis & site directed mutagenesisAnuKiruthika
This document provides information on transposon mutagenesis and site-directed mutagenesis. It defines transposons as "jumping genes" that can move locations in the genome and explains that transposon mutagenesis uses transposons to cause mutations by interrupting genes. Site-directed mutagenesis is described as an in vitro technique to introduce specific mutations into DNA using methods like conventional PCR, nested PCR, or inverse PCR. The document outlines several applications of these mutagenesis methods such as identifying virulence genes or screening for desired mutations.
1) Germplasm conservation involves preserving genetic material, such as seeds, cells, tissues, and body parts, through in-situ and ex-situ methods to maintain biodiversity and provide resources for breeding programs.
2) Cryopreservation at ultra-low temperatures in liquid nitrogen is an important ex-situ technique that can preserve germplasm long-term without subculturing. It involves preculturing plant materials, treating with cryoprotectants, and either slow-freezing or vitrification prior to storage in liquid nitrogen.
3) A case study demonstrates the successful cryopreservation of mint shoot tips using encapsulation-dehydration and PVS2-vitrification, with
Agrobacterium tumefaciens is a soil bacterium that causes crown gall disease in plants. It does this by transferring a segment of its tumor-inducing plasmid (Ti plasmid) called T-DNA into the plant's DNA. The T-DNA contains genes that cause uncontrolled cell growth. Researchers developed techniques using the Ti plasmid and Agrobacterium to genetically transform plants by replacing the tumor-causing genes with desired genes. This involves either a binary vector system with the T-DNA on a separate small plasmid, or co-integration of a new plasmid containing the gene of interest into the Ti plasmid. Transformed plants can be regenerated from infected plant cells or tissues.
This document discusses protoplast isolation and fusion. It defines a protoplast as a plant, bacterial, or fungal cell that has had its cell wall removed, leaving the plasma membrane intact. The document outlines the history of protoplast isolation, from the first isolation in 1892 to the use of commercial enzyme mixtures in the 1960s. It also describes how protoplasts can be fused using polyethylene glycol or other fusogens to create somatic hybrids for plant breeding purposes.
Genetic engineering for abiotic stress toleranceSachin Ekatpure
This document discusses various approaches for improving plant tolerance to abiotic stresses through transgenic methods. It summarizes 7 approaches: 1) Engineering genes for osmolyte biosynthesis like proline and glycine betaine, 2) Engineering genes encoding enzymes that scavenge reactive oxygen species, 3) Engineering genes encoding LEA proteins, 4) Engineering genes encoding enzymes with different temperature optima, 5) Engineering molecular chaperone genes, 6) Engineering transcription factor genes, and 7) Engineering plant cell membrane genes. For each approach, it provides examples of transgenic plants that were developed and their improved stress tolerance performances.
This document discusses breeding for resistance to abiotic stresses like drought, salt, and cold in fruit crops. It provides information on the characteristics, effects, and mechanisms of different abiotic stresses. It also outlines strategies for breeding resistance, including selecting from cultivated varieties, landraces, and wild relatives. The key mechanisms of resistance include avoidance, tolerance, and acclimation. Traits like early maturity, reduced transpiration, and accumulating osmolytes can provide drought and salt resistance.
In-planta transformation is a direct transformation method that does not require tissue culture steps. It has several advantages over traditional transformation techniques, including being more efficient, quick, and not requiring regeneration systems. Various in-planta methods have been developed, including floral dip, vacuum infiltration, and pollen-tube mediated transformation. However, transformation efficiencies tend to be low across methods and plant species. Further optimization is still needed to improve transformation frequencies and consistency of transgene expression for many crop species using in-planta transformation approaches.
A process where an embryo is derived from a single somatic cell or group of somatic cells. Somatic embryos (SEs) are formed from plant cells that are not normally involved in embryo formation.
Embryos formed by somatic embryogenesis are called Embryoids.
The process was discovered for the first time in Daucas carota L. (carrot) by Steward (1958), Reinert (1959).
Transfer and establishment of whole plant in soilmadgenius368
This document discusses the process of acclimatizing micropropagated plantlets to soil conditions. It explains that plantlets grown in vitro have physiological differences from soil-grown plants. The acclimatization process aims to increase plantlet growth and survival rates through controlling the physical and chemical environment. Some key steps discussed are hardening plantlets to different light and humidity levels gradually, supplying carbohydrates, using plant growth retardants, and exposing plantlets to beneficial microbes to prepare them for biotic stresses in soil. The document also mentions some alternative and low-cost methods for acclimatization using different substratum.
Plant regeneration is possible through organogenesis or somatic embryogenesis where differentiated plant cells can become totipotent using hormones. The process involves taking an explant from a plant and culturing it in a nutrient medium under sterile conditions to regenerate a whole new plant. Somaclonal variation may occur during this process, introducing genetic changes into the regenerated plants.
Agrobacterium mediated gene transfer in plants.ICHHA PURAK
This power point presentation consist of 41 slides. Attempts have been made to illustrate how Agrobacterium behaves us natural genetic engineer. How it can infect a plant through wound and a part of DNA present on Ti plasmid is Tranferred and causes disease as crown gall in the infected plant. In second part of the presentation attempts have been made to describe how Agrobacterium can be utilized for iinsertion of desired gene into the plant,what manipulation are to be made with Agrobacterium.How infection and transfer of desired gene can be made possible.What is the role of plant tissue culture etc.
The document summarizes the mechanism of T-DNA transfer during Agrobacterium tumefaciens infection. It explains that T-DNA is a fragment of DNA transferred from the tumor-inducing (Ti) plasmid of A. tumefaciens into the host plant genome. The T-DNA is bordered by repeats and encodes genes that cause tumors in the plant. Virulence genes are expressed in response to plant signals and produce single-stranded T-DNA, which forms a complex with other proteins and is transported into the plant cell and integrated into the plant nuclear DNA, causing uncontrolled cell growth and tumor formation. The mechanism involves multiple virulence protein complexes and integration of T-DNA is directed by the
Chloroplasts are double-membrane organelles found in plant cells that contain chlorophyll and are the site of photosynthesis. Chloroplast DNA is circular and ranges in size from 120,000 to 170,000 base pairs. It contains approximately 120 genes, including genes that encode proteins involved in photosynthesis and the transcription and translation machinery. Chloroplast DNA replication is semi-conservative and there are typically multiple copies of the chloroplast genome within each chloroplast.
An overview of the Agrobacterium-mediated gene transfer process. Moreover, studied different kinds of Agrobacterium species are involved in this mechanism.
Agrobacterium is a rod-shaped, Gram-negative bacteria found mostly in the soil. It is a plant pathogen that is responsible for causing crown gall disease in them. This bacteria is also known as the natural genetic engineer because of it's the ability to integrate its plasmid Gene into the plant genome.
Agrobacterium tumefaciens transfer of their genetic material T-DNA of Ti-plasmid into the plant cell: A: Agrobacterium tumefaciens; B: Agrobacterium genome; C: Ti Plasmid : a: T-DNA , b: Vir genes , c: Replication origin , d: Opines catabolism genes; D: Plant cell
A Ti-Plasmid (tumor-inducing plasmid) is a ds, circular DNA that often, but not always. It's a piece of genetic equipment that transfers genetic material from bacterial cells means Agrobacterium tumefaciens into plant cells used to induce tumors in the plant. The Ti-plasmid is damage when Agrobacterium is grown above 28 °C. Such cured bacteria don't induce crown gall disease in the plant due to they are avirulent. The Ti-Plasmid are classified into two types on the basis of opine genes are present in T-DNA.
The Plasmid has 196 genes that code for 195 proteins. There is no one structural RNA. The plasmid is 206.479 nucleotides long. the GC content is 56% and 81% of the genetic material is coding genes.
The modification of this plasmid is a very important source in the production of transgenic plants.
The T-DNA must be cut out of the circular plasmid. A VirD1/D2 complex nicks the DNA at the left and right border sequences. The VirD2 protein is covalently attached to the 5' end. VirD2 contains a motif that leads to the nucleoprotein complex being targeted to the type IV secretion system (T4SS).
In the cytoplasm of the recipient cell, the T-DNA complex becomes coated with VirE2 proteins, which are exported through the T4SS independently from the T-DNA complex. Nuclear localization signals, or NLS, located on the VirE2 and VirD2 are recognized by the importin alpha protein, which then associates with importin beta and the nuclear pore complex to transfer the T-DNA into the nucleus. So that the T-DNA can integrate into the host genome.
We inoculate Agrobacterium containing our genes of interest, onto wounded plant tissue explants. The Agrobacterium then transfers the gene of interest into the DNA of the plant tissue.
Auxins biosynthesis physiological role and mechanismpavanknaik
Auxins are plant hormones that regulate growth and development. The main auxin is indole-3-acetic acid (IAA) which is synthesized from the amino acid tryptophan through several pathways. IAA is transported from shoot tips to regions of elongation through active transport and influences growth through effects on cell wall plasticity and gene expression. Auxins have many physiological roles including stem elongation, apical dominance, root initiation, fruit development, and growth responses to light.
Plant disease resistance occurs through both pre-formed structures and infection-induced immune responses. There are two tiers of the plant immune system - pattern-triggered immunity (PTI) triggered by pathogen-associated molecular patterns (PAMPs), and effector-triggered immunity (ETI) triggered by recognition of pathogen effectors through resistance (R) proteins. Quantitative resistance involving multiple genes provides more durable resistance than major gene resistance. Genetic engineering and breeding can enhance crop disease resistance through introduction of R genes or resistance mechanisms.
Genetic engineering can be used to develop drought tolerant crops. Approaches include engineering genes for organic osmolytes like glycine betaine, trehalose, proline, and mannitol. Genes encoding regulatory enzymes and LEA proteins can also be engineered for drought tolerance. Transgenic crops overexpressing antioxidant enzymes or transcription factors involved in the plant stress response have shown enhanced drought tolerance. Commercially available drought tolerant crops include Monsanto's DroughtGard maize which expresses a cold shock protein from bacteria.
The document discusses Agrobacterium rhizogenes, which causes hairy root disease in plants. It contains genes on Ri plasmids, including rol genes, that are transferred to plant cells and cause root proliferation. The T-DNA from the plasmid is integrated into the plant genome. Agrobacterium attaches to wounded plant cells, vir genes are activated by compounds from the plant, and the T-DNA and virulence genes work together to transfer T-DNA to the plant cell. Hairy roots can be grown in vitro for research on plant-pathogen interactions and production of compounds.
This document discusses germplasm and its conservation. It begins by defining germplasm as a collection of genetic resources for an organism, such as a seed bank or gene bank, that contains the genetic information for a species. Germplasm conservation is important to preserve genetic diversity and provide plant breeders resources to develop new crop varieties. Methods of conservation include in situ conservation of plants in their natural habitat and ex situ conservation of seeds, tissues, cells or DNA stored outside the natural habitat. Cryopreservation in liquid nitrogen at -196°C is an effective long-term storage method that stops cellular metabolism. The document outlines the cryopreservation process and applications for conserving plant species and genetic variations.
This document discusses somaclonal variation, which refers to genetic variation that arises during tissue culture or plant regeneration from cell cultures. It provides definitions and history of the term as coined by Larkin and Scowcroft in 1981. The document outlines the various causes and types of somaclonal variation including physiological, genetic, and biochemical causes. It also describes methods for generating somaclonal variation both with and without in vitro selection. Finally, it discusses applications for detecting and isolating somaclonal variants, particularly for developing disease resistance in various crop species.
This document discusses genetically modified Golden Rice and arguments for allowing its use. It begins with an overview of the Allow Golden Rice Now! campaign and its goals of education and protest. It then provides background on plant breeding and defines relevant terms. The rest of the document outlines different breeding techniques, organizations that support Golden Rice's safety, its potential benefits in addressing vitamin A deficiency, and Greenpeace's opposition to it through destruction of field trials and claims not supported by scientific evidence.
Vitamin A deficiency (VAD) can cause xerophthalmia, a disease affecting the eyes. Early signs include night blindness and dryness of the conjunctiva. More severe forms include corneal lesions that can lead to blindness. VAD is most common in young children in developing countries where diets lack vitamin A-rich foods. Treatment involves high doses of vitamin A supplements to restore levels as well as addressing underlying malnutrition. Prevention focuses on dietary diversification, supplementation programs, and promoting breastfeeding.
1) Germplasm conservation involves preserving genetic material, such as seeds, cells, tissues, and body parts, through in-situ and ex-situ methods to maintain biodiversity and provide resources for breeding programs.
2) Cryopreservation at ultra-low temperatures in liquid nitrogen is an important ex-situ technique that can preserve germplasm long-term without subculturing. It involves preculturing plant materials, treating with cryoprotectants, and either slow-freezing or vitrification prior to storage in liquid nitrogen.
3) A case study demonstrates the successful cryopreservation of mint shoot tips using encapsulation-dehydration and PVS2-vitrification, with
Agrobacterium tumefaciens is a soil bacterium that causes crown gall disease in plants. It does this by transferring a segment of its tumor-inducing plasmid (Ti plasmid) called T-DNA into the plant's DNA. The T-DNA contains genes that cause uncontrolled cell growth. Researchers developed techniques using the Ti plasmid and Agrobacterium to genetically transform plants by replacing the tumor-causing genes with desired genes. This involves either a binary vector system with the T-DNA on a separate small plasmid, or co-integration of a new plasmid containing the gene of interest into the Ti plasmid. Transformed plants can be regenerated from infected plant cells or tissues.
This document discusses protoplast isolation and fusion. It defines a protoplast as a plant, bacterial, or fungal cell that has had its cell wall removed, leaving the plasma membrane intact. The document outlines the history of protoplast isolation, from the first isolation in 1892 to the use of commercial enzyme mixtures in the 1960s. It also describes how protoplasts can be fused using polyethylene glycol or other fusogens to create somatic hybrids for plant breeding purposes.
Genetic engineering for abiotic stress toleranceSachin Ekatpure
This document discusses various approaches for improving plant tolerance to abiotic stresses through transgenic methods. It summarizes 7 approaches: 1) Engineering genes for osmolyte biosynthesis like proline and glycine betaine, 2) Engineering genes encoding enzymes that scavenge reactive oxygen species, 3) Engineering genes encoding LEA proteins, 4) Engineering genes encoding enzymes with different temperature optima, 5) Engineering molecular chaperone genes, 6) Engineering transcription factor genes, and 7) Engineering plant cell membrane genes. For each approach, it provides examples of transgenic plants that were developed and their improved stress tolerance performances.
This document discusses breeding for resistance to abiotic stresses like drought, salt, and cold in fruit crops. It provides information on the characteristics, effects, and mechanisms of different abiotic stresses. It also outlines strategies for breeding resistance, including selecting from cultivated varieties, landraces, and wild relatives. The key mechanisms of resistance include avoidance, tolerance, and acclimation. Traits like early maturity, reduced transpiration, and accumulating osmolytes can provide drought and salt resistance.
In-planta transformation is a direct transformation method that does not require tissue culture steps. It has several advantages over traditional transformation techniques, including being more efficient, quick, and not requiring regeneration systems. Various in-planta methods have been developed, including floral dip, vacuum infiltration, and pollen-tube mediated transformation. However, transformation efficiencies tend to be low across methods and plant species. Further optimization is still needed to improve transformation frequencies and consistency of transgene expression for many crop species using in-planta transformation approaches.
A process where an embryo is derived from a single somatic cell or group of somatic cells. Somatic embryos (SEs) are formed from plant cells that are not normally involved in embryo formation.
Embryos formed by somatic embryogenesis are called Embryoids.
The process was discovered for the first time in Daucas carota L. (carrot) by Steward (1958), Reinert (1959).
Transfer and establishment of whole plant in soilmadgenius368
This document discusses the process of acclimatizing micropropagated plantlets to soil conditions. It explains that plantlets grown in vitro have physiological differences from soil-grown plants. The acclimatization process aims to increase plantlet growth and survival rates through controlling the physical and chemical environment. Some key steps discussed are hardening plantlets to different light and humidity levels gradually, supplying carbohydrates, using plant growth retardants, and exposing plantlets to beneficial microbes to prepare them for biotic stresses in soil. The document also mentions some alternative and low-cost methods for acclimatization using different substratum.
Plant regeneration is possible through organogenesis or somatic embryogenesis where differentiated plant cells can become totipotent using hormones. The process involves taking an explant from a plant and culturing it in a nutrient medium under sterile conditions to regenerate a whole new plant. Somaclonal variation may occur during this process, introducing genetic changes into the regenerated plants.
Agrobacterium mediated gene transfer in plants.ICHHA PURAK
This power point presentation consist of 41 slides. Attempts have been made to illustrate how Agrobacterium behaves us natural genetic engineer. How it can infect a plant through wound and a part of DNA present on Ti plasmid is Tranferred and causes disease as crown gall in the infected plant. In second part of the presentation attempts have been made to describe how Agrobacterium can be utilized for iinsertion of desired gene into the plant,what manipulation are to be made with Agrobacterium.How infection and transfer of desired gene can be made possible.What is the role of plant tissue culture etc.
The document summarizes the mechanism of T-DNA transfer during Agrobacterium tumefaciens infection. It explains that T-DNA is a fragment of DNA transferred from the tumor-inducing (Ti) plasmid of A. tumefaciens into the host plant genome. The T-DNA is bordered by repeats and encodes genes that cause tumors in the plant. Virulence genes are expressed in response to plant signals and produce single-stranded T-DNA, which forms a complex with other proteins and is transported into the plant cell and integrated into the plant nuclear DNA, causing uncontrolled cell growth and tumor formation. The mechanism involves multiple virulence protein complexes and integration of T-DNA is directed by the
Chloroplasts are double-membrane organelles found in plant cells that contain chlorophyll and are the site of photosynthesis. Chloroplast DNA is circular and ranges in size from 120,000 to 170,000 base pairs. It contains approximately 120 genes, including genes that encode proteins involved in photosynthesis and the transcription and translation machinery. Chloroplast DNA replication is semi-conservative and there are typically multiple copies of the chloroplast genome within each chloroplast.
An overview of the Agrobacterium-mediated gene transfer process. Moreover, studied different kinds of Agrobacterium species are involved in this mechanism.
Agrobacterium is a rod-shaped, Gram-negative bacteria found mostly in the soil. It is a plant pathogen that is responsible for causing crown gall disease in them. This bacteria is also known as the natural genetic engineer because of it's the ability to integrate its plasmid Gene into the plant genome.
Agrobacterium tumefaciens transfer of their genetic material T-DNA of Ti-plasmid into the plant cell: A: Agrobacterium tumefaciens; B: Agrobacterium genome; C: Ti Plasmid : a: T-DNA , b: Vir genes , c: Replication origin , d: Opines catabolism genes; D: Plant cell
A Ti-Plasmid (tumor-inducing plasmid) is a ds, circular DNA that often, but not always. It's a piece of genetic equipment that transfers genetic material from bacterial cells means Agrobacterium tumefaciens into plant cells used to induce tumors in the plant. The Ti-plasmid is damage when Agrobacterium is grown above 28 °C. Such cured bacteria don't induce crown gall disease in the plant due to they are avirulent. The Ti-Plasmid are classified into two types on the basis of opine genes are present in T-DNA.
The Plasmid has 196 genes that code for 195 proteins. There is no one structural RNA. The plasmid is 206.479 nucleotides long. the GC content is 56% and 81% of the genetic material is coding genes.
The modification of this plasmid is a very important source in the production of transgenic plants.
The T-DNA must be cut out of the circular plasmid. A VirD1/D2 complex nicks the DNA at the left and right border sequences. The VirD2 protein is covalently attached to the 5' end. VirD2 contains a motif that leads to the nucleoprotein complex being targeted to the type IV secretion system (T4SS).
In the cytoplasm of the recipient cell, the T-DNA complex becomes coated with VirE2 proteins, which are exported through the T4SS independently from the T-DNA complex. Nuclear localization signals, or NLS, located on the VirE2 and VirD2 are recognized by the importin alpha protein, which then associates with importin beta and the nuclear pore complex to transfer the T-DNA into the nucleus. So that the T-DNA can integrate into the host genome.
We inoculate Agrobacterium containing our genes of interest, onto wounded plant tissue explants. The Agrobacterium then transfers the gene of interest into the DNA of the plant tissue.
Auxins biosynthesis physiological role and mechanismpavanknaik
Auxins are plant hormones that regulate growth and development. The main auxin is indole-3-acetic acid (IAA) which is synthesized from the amino acid tryptophan through several pathways. IAA is transported from shoot tips to regions of elongation through active transport and influences growth through effects on cell wall plasticity and gene expression. Auxins have many physiological roles including stem elongation, apical dominance, root initiation, fruit development, and growth responses to light.
Plant disease resistance occurs through both pre-formed structures and infection-induced immune responses. There are two tiers of the plant immune system - pattern-triggered immunity (PTI) triggered by pathogen-associated molecular patterns (PAMPs), and effector-triggered immunity (ETI) triggered by recognition of pathogen effectors through resistance (R) proteins. Quantitative resistance involving multiple genes provides more durable resistance than major gene resistance. Genetic engineering and breeding can enhance crop disease resistance through introduction of R genes or resistance mechanisms.
Genetic engineering can be used to develop drought tolerant crops. Approaches include engineering genes for organic osmolytes like glycine betaine, trehalose, proline, and mannitol. Genes encoding regulatory enzymes and LEA proteins can also be engineered for drought tolerance. Transgenic crops overexpressing antioxidant enzymes or transcription factors involved in the plant stress response have shown enhanced drought tolerance. Commercially available drought tolerant crops include Monsanto's DroughtGard maize which expresses a cold shock protein from bacteria.
The document discusses Agrobacterium rhizogenes, which causes hairy root disease in plants. It contains genes on Ri plasmids, including rol genes, that are transferred to plant cells and cause root proliferation. The T-DNA from the plasmid is integrated into the plant genome. Agrobacterium attaches to wounded plant cells, vir genes are activated by compounds from the plant, and the T-DNA and virulence genes work together to transfer T-DNA to the plant cell. Hairy roots can be grown in vitro for research on plant-pathogen interactions and production of compounds.
This document discusses germplasm and its conservation. It begins by defining germplasm as a collection of genetic resources for an organism, such as a seed bank or gene bank, that contains the genetic information for a species. Germplasm conservation is important to preserve genetic diversity and provide plant breeders resources to develop new crop varieties. Methods of conservation include in situ conservation of plants in their natural habitat and ex situ conservation of seeds, tissues, cells or DNA stored outside the natural habitat. Cryopreservation in liquid nitrogen at -196°C is an effective long-term storage method that stops cellular metabolism. The document outlines the cryopreservation process and applications for conserving plant species and genetic variations.
This document discusses somaclonal variation, which refers to genetic variation that arises during tissue culture or plant regeneration from cell cultures. It provides definitions and history of the term as coined by Larkin and Scowcroft in 1981. The document outlines the various causes and types of somaclonal variation including physiological, genetic, and biochemical causes. It also describes methods for generating somaclonal variation both with and without in vitro selection. Finally, it discusses applications for detecting and isolating somaclonal variants, particularly for developing disease resistance in various crop species.
This document discusses genetically modified Golden Rice and arguments for allowing its use. It begins with an overview of the Allow Golden Rice Now! campaign and its goals of education and protest. It then provides background on plant breeding and defines relevant terms. The rest of the document outlines different breeding techniques, organizations that support Golden Rice's safety, its potential benefits in addressing vitamin A deficiency, and Greenpeace's opposition to it through destruction of field trials and claims not supported by scientific evidence.
Vitamin A deficiency (VAD) can cause xerophthalmia, a disease affecting the eyes. Early signs include night blindness and dryness of the conjunctiva. More severe forms include corneal lesions that can lead to blindness. VAD is most common in young children in developing countries where diets lack vitamin A-rich foods. Treatment involves high doses of vitamin A supplements to restore levels as well as addressing underlying malnutrition. Prevention focuses on dietary diversification, supplementation programs, and promoting breastfeeding.
Golden Rice is a genetically engineered rice variety that produces beta-carotene, which the body converts to vitamin A. It was developed to help address vitamin A deficiency in developing countries. The Golden Rice Project inserted genes for phytoene synthase and lycopene cyclase enzymes into rice, allowing it to synthesize beta-carotene in the edible endosperm. Golden Rice 2 produces 23 times more carotenoids than the original. Potential advantages include providing a sustainable source of vitamin A, but there are also concerns about allergies, expression levels, and impacts on biodiversity and culture. Further research and development is still needed before Golden Rice can be effectively deployed.
Golden rice was developed to address vitamin A deficiency in rice-dependent populations. It contains genes from maize and soil bacteria that produce beta-carotene, a vitamin A precursor. Over 400 million people worldwide are vitamin A deficient due to their reliance on rice, which lacks vitamin A and other micronutrients. Golden rice aims to biofortify rice to help prevent blindness and deaths from vitamin A deficiency.
Golden Rice – A Humanitarian Biotechnology Projectsol777
Presentation of Golden Rice Co-inventor Peter Beyer, professor at the university of Freiburg, at the Comm4Biotech conference 2011 in Strasbourg - more information on http://www.comm4biotech.eu .
The document discusses several major genetically modified crops including golden rice, GM soybeans, GM tomatoes, and GM corn. Golden rice has been engineered to produce beta-carotene which provides vitamin A. GM soybeans are mostly herbicide resistant and used for livestock feed. The first GM food available was the FlavrSavr tomato, which had longer shelf life. Bt corn produces its own pesticide while herbicide tolerant corn is resistant to weed killers.
Transgenic plant with improved nutritional qualityDr. Kirti Mehta
This document summarizes the development of Golden Rice, a genetically engineered rice variety that produces beta-carotene, a precursor of vitamin A. It was developed to address vitamin A deficiency in developing countries where rice is a staple crop. The document describes how researchers introduced genes from daffodil and bacteria to complete the beta-carotene biosynthesis pathway in rice endosperm. Early research demonstrated beta-carotene production in transgenic rice. Further work improved beta-carotene levels and introduced the trait into indica rice varieties commonly consumed in Asia where vitamin A deficiency is widespread. The goal of Golden Rice is to provide a sustainable solution to prevent blindness and other health issues caused by vitamin A deficiency.
The document discusses transgenic potential and provides examples of second generation genetically modified crops. It summarizes the traits and benefits of non-browning apples, non-bruising and low acrylamide potatoes, and maize varieties with low phytic acid and increased essential amino acids. It also discusses healthier oils from soybean and canola that have modified fatty acid compositions. The document then provides background information on techniques used to modify these crops, such as gene silencing and adding or modifying specific genes.
modification of flower pigmentation & nutritional content ppt for students a...thirupathiSathya
To improve flower appearance and postharvest lifetime.
By traditional breeding techniques thousands of new varieties that differ from one another in color, shape, and plant architecture has been created.
But this is a slow and pains taking procedure
Uniquely colored flowers can be developed by manipulating the genes for enzymes in the anthocyanin biosynthesis pathway.
Anthocyanin is a flavonoids common type of flower pigmentation.
Four plants
Roses
Carnations
Tulips
Chrysanthemums
Used to form unique colours.
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This document discusses the application of recombinant DNA technology to improve seed storage proteins and develop Golden Rice. It provides background on transgenic plants and their benefits, including improved traits, yields, and stress resistance. It describes seed storage proteins and their importance as a protein source. Golden Rice is introduced as an example of a transgenic crop developed to provide vitamin A through the addition of two beta-carotene biosynthesis genes to rice. The document covers the process used to develop Golden Rice and its benefits and limitations for addressing vitamin A deficiency.
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1. Golden Rice –
A Humanitarian Biotechnology Project
COMM4BIOTECH Congress; 18-19 Nov. 2011
Peter Beyer, Center for Applied Biosciences, University of Freiburg, Germany
2. WHY?
Population growth
10000
8000
Population [million]
6000
4000
Developing Nations
2000
0
Industrialized Nations
1990 2000 2010 2020 2030 2040 2050
Year Source: C. Lupi, BATS report, I/95
3. Directly, or indirectly, plants provide all of
humanity’s food
During the past 100 years agriculture has (of necessity)
focused on increasing yield. Nutrient content has
largely been ignored in breeding.
More of our food comes from fewer species (54% from
corn, rice and wheat). We’ve lost biochemical diversity
in the staples in our diets.
Plants, in principle provide all macro -and
micronurients required, but the latter are very
unevenly distributed.
4. Share of daily energy intake
for rural Bangladesh
Staples (rice)
Non-staples
(Vegetables)
84 %
Fish and other
animal sources
Source: H. Bouis, IFPRI; HarvestPlus
5. Directly, or indirectly, plants provide all of
humanity’s food
During the past 100 years agriculture has (of necessity)
focused on increasing yield. Nutrient content has
largely been ignored in breeding.
More of our food comes from fewer species (54% from
corn, rice and wheat). We’ve lost biochemical diversity
in our diets.
Plants provide all macro -and micronurients required,
but the latter are very unevenly distributed in plant
tissues.
7. Nutritional Diversity
Iron, Zinc Folate Provit A Vit. E
Facts:
Rice Three billion live on-less than - $ per day,
- 2 -
1.5 billion on less than 1 $ per day and cannot afford a
Tomato - - (+) +
diversified diet or industrially produced supplements
Beans + + - +
Spinach + + + +
Meat + + Vit A + -
• Millions are chronically micronutrient
malnourished
8. Intervention strategies:
Supplementation
Industrial fortification
Education
All necessary and very valuable but
there are drawbacks:
Distrubution, educated medical staff
Centrally processed food items
Only partially applicable
Economically sustainable?
9. Biofortification is a cost effective alternative to
classical interventions
Improve the nutritional value of agronomically important crop
tissues through
Breeding
Recombinant DNA technology
is all achievable through breeding ?
Simple answer: NO!!
Some crops cannot be bred or breeding is very difficult
Trait variability is insufficient
www.harvestplus.org
10. Some traits cannot be bred because there is no
adequate trait variability
Rice, (polished grains) for instance
Provitamin A: No „yellow grains“ in
germplasm collections
Folate: Practically absent
Iron: low variability, ranging from 1 – 8 ppm (Target 14 ppm)
Zinc: much more important variability, ranging from 16 – 28 ppm
(Target: 24 ppm)
11. Golden Rice cannot be bred
….the application of recombinant
DNA technology is necessary
Breeding where possible
Genetic modification where necessary
12. Why engineering ß-carotene
(provitamin A) biosynthesis into rice
endosperm?
Milled rice is provitamin-a-free
Symptoms of a provitamin-a-free diet
• Night-blindness
• Xerophthalmia
• Fatal susceptibility to childhood diseases (e.g. measles) and
general infections (diarrhoea, respiratory diseases)
Epidemiology
• 124 million children are deficient in vitamin A
• 1-2 million deaths annually (1-4 years)
• 0.25-0.5 million deaths (5-10 years) UNICEF; Humphrey et al.,
1992)
• A severe public health problem in (118) countries (WHO)
14. Why engineering ß-carotene
(provitamin A) biosynthesis into rice
endosperm?
Milled rice is provitamin-a-free
Symptoms of a provitamin-a-free diet
• Night-blindness
• Xerophthalmia
• Fatal susceptibility to childhood diseases (e.g. measles) and
general infections (diarrhoea, respiratory diseases)
Epidemiology
• 124 million children are deficient in vitamin A
• 1-2 million deaths annually (1-4 years)
• 0.25-0.5 million deaths (5-10 years)
• A severe public health problem in (118) countries (WHO)
15. Global prevalence of vitamin A deficiency in populations at risk 1995–2005
WHO Global Database on Vitamin A Deficiency
25. Prototypes:
Not apt for
product development
Construct ill-defined
Integration ill-defined
Antibiotic selectable marker
Low amount of bC (1.6 µg/g)
Start from scratch
include Indica rice
varieties.
Happy Easter
Ye et al., 2000; Science 287:303
26. Improved Golden Rice variants came in two versions
In the public and in the private sector
Gt1p PSY (Np) Gt1p tp-CrtI
(from Narcissus)
No selectable marker gene
Ca. 1000 events
„Clean“ integration selected
CrtI controlled by an endosperm-specific promoter
Three high expressing events preselected
Known as Golden Rice 1
Technology works in Indica varieties
29. Improvements:
Five years of research
were dominated by
efforts to increase the
amount of provitamin
A in GR both, in the
Public Sector as well
as at Syngenta
31. Phytoene synthase was investigated by Rachel Drake (Syngenta)
Because PSY expression is good in GR, different versions of the
PSY gene were assayed.
Seed promoter CrtI Seed promoter Daffodil Psy Ubi promoter hygR
Maize Psy
Rice Psy
Tomato Psy
Transformation into a japonica short-grain rice,
(Asanohikare) 20+ plants each Pepper Psy
20
18
Rice and Maize PSY (E3)
Carotenoid content (μg g dwt)
16
14
best. Proportion of
-1
12
10
8
ß-carotene increased.
6
4
2
0
rice maize pepper tomato daffodil
Psy/crtI Psy/crtI Psy/crtI Psy/crtI Psy/crtI
Individual transgenic plant (event) Paine et al., 2005 Nat Biotechnol. 23, 482-487
32. E3 (PSY is rate-limiting)
OK
OK
OK OK
Too slow!!!
E8
CrtI E1
CrtI CrtI E3 E2
Precursor
Zwischenprodukt
Produkt
33. Golden Rice 2 was re-made for implementation
GT1pI tp-CrtI GT1pI ZmPSY ubi1p PMI
pSYN12424
Transform long grain rice variety (Kaybonnet)
Sugar selectable marker (PMI)
619 individual GM rice plants
Screen for seed colour, gene copy number, fertility
Select 6 “Golden Rice 2” events for
further screening and development
34. Improved provitamin A
accumulation in Golden
Rice I and II
Increase in provit-A content: ca. 10-fold over GR1; about 25-fold
over the prototype
35. GR was a breeding project during the past 4 years
9 events into 15 selected varieties (MAS), preceding event selection
IR64 & IR36: Mega-varieties with broad Asian coverage
BRRI dhan 29 : The most popular boro rice variety in Bangladesh
PSB Rc82: The most popular rice variety in the Philippines
OS 6561: Most popular in Vietnam
Chehirang: Leading variety in Indonesia (with IRRC)
Stacking With other micronutrients (zinc, iron vit E, lysine)
Stacking With new agronomic traits (Submergence tolerance)
Partner Institutions:
o IRRI (Int.), lead
o Philrice (Philippines)
o BRRI (Bangladesh)
o CLRRI (Vietnam)
o IARI (India)
o TNAU (India)
o DRR (India)
o Huazhong Univ. (China)
36. Event selection (All single locus intact ingtegration)
Which event(s) produce consistent levels of povitamin A across cultivars?
Which event(s) reproduce consistently the characteristics of the recurrent
parents? (Completed, all other events destroyed)
The big question:
Human bioavailability study
Which level of provitamin A must be delivered by GR to be effective?
Conducted at TUFTS first and then in China
Bioconversion determined: 3.8-to-1 (!!!)
Tang et al., (2009) Golden Rice is an effective source of vitamin A. Am J Clin Nutr89:1776–83
37. Event selection (All single locus intact ingtegration)
Which event(s) produce consistent levels of povitamin A across cultivars?
Which event(s) reproduce consistently the characteristics of the recurrent
parents? (Completed, all other events destroyed)
The big question:
All done?
Which level of provitamin A must be delivered by GR to be effective?
Human bioavailability study
Conducted at TUFTS first and then in China
Bioconversion determined: 3.8-to-1 (!!!)
Tang et al., (2009) Golden Rice is an effective source of vitamin A. Am J Clin Nutr89:1776–83
38. Regulatory dossiers can be complex and expensive
Event independent studies
Exposure evaluation
Modelling analysis for intended use.
Bioavailability study.
Protein production and equivalence
Extraction from GMO plant or heterologous source
Biochemical characterisation
Function/ specificity/ mode of action.
Protein evaluation
No homology with toxins and allergens.
Rapid degredation in gastric /intestinal studies.
Heat lability
No indication of acute toxicity in rodents.
Further allergenicity assessments
39. Event dependent studies
Molecular characterization and genetic stability
Single copy effect; marker gene at same locus.
Simple integration; Mendelian inheritance over
three generations (minimum).
No potential gene disruption.
No unknown open reading frames.
No DNA transfer beyond borders.
No antibiotic resistance gene or origin of replication.
Insert limited to the minimum necessary.
Insert plus flanking plant genome sequenced.
Phenotypic evidence for stability over 3 generations
Biochemical evidence for stability.
Unique DNA identifier for tracebility/detection.
Expression profiling
Gene expression levels at key growth stages.
Evidence for seed-specific expression.
40. Phenotype analysis
Field performance, typical agronomic traits, yield -
compared to isogenic lines.
Pest and disease status to be same as isogenic background.
Compositional analysis
Data from 2 seasons x 6 locations x 3 reps. on proximates, macro and micro
nutrients, antinutrients, inherent toxins and allergens. Data generated on
modified and isogenic background.
Environmental risk assessment
Minimize potential for gene flow.
Evaluate any change in insect preference – by field survey.
Data submitted must be of scientific publication quality
(but will seldomly result in academic recognition)
Significant Costs
41. Why in the „Public Sector“?
Some products of high general demand are not commercialy interesting.
The private sector will not make major investments. Hence the public sector
should take over.
Funding is required
To conduct more proof-of principle experimentation and translational
research
No expectations for funding in €
Public research funding on transgenic crops in Europe goes into
„Biosafety“ research. There is virtually no funding for research that goes
beyond discovery.
(the development if GR relies on US-Philanthropy and on National funding in
GR-implementing countries )
42. The „Danger Zone“ is in Europe
Requirements:
Raised to the point where anything that appears technically
feasible is being requested/offered to be applied. This is in
part driven by some public sector scientists.
Justification:
Genetically modified plants are little understood (while we
understand traditionally bred or mutagenized varieties?)
Kuiper et al., 2001 Assessment of the food safety issues related to
genetically modified foods. Plant J. 2001 Sep;27(6):503-28.
Consequence:
Can raise costs to unaffordable levels - delays.
43. Overregulation of novel technologies: the „Locomotive Act“
England, 1861 (the Red Flag Act)
Self-propelled vehicles on public roads in the
United Kingdom must be preceded by a man
on foot waving a red flag and
blowing a horn.
This effectively killed road auto development
in the UK for most of the rest of the 19th
century
The red flag law was not
repealed until 1896
44. Where we are in the timeline?
(Philippines - first launch)
Production of stable lines; fingerprinting
2007 (Bioavailability, biosafety & other studies)
2008 BPI application
Single location trial of elite lines (2
2009 seasons), generation of protein data
Multi-location trials; gather
2010 agronomic and biosafety data
(2 seasons)
2013/14 Release
The Philippines has experience in the regulatory review of a transgenic nutritionally-
enhanced crop: LY038. 7 GMOs approved in the Philippines (http://agbios.com/dbase.php)
45. First Outdoor Trial
of GR in Asia
IR64 GR1 event 309; 20 lines
Transplanting at IRRI on April 2, 2008 May 30, 2008
April 10, 2008
46. Activities beyond technical and safety issues
Golden Rice:
communication & social marketing
Consumers
Growers
(+ Policy Makers, Heath Community, Regulators, Traders, NGOs, etc. )
Focus groups: Qualitative information on consumer &
grower attitudes in vitamin A-deficient areas
47. The Focus Group
normally 10-12 people for structured open ended discussion for
2 hours, with careful recording
MBA students of the Asian Institute of Management,
Manila –training at AIM, IRRI. PhilRice
48. First social marketing research was coducted in 2009
(then continued by a company)
Focus Group Methodology
In 4 selected islands in the Philippines
AIM-MBA students teams from these
areas
~ 720 hours interviews with 360
individuals in selected groups: gender,
rice farming or not farming
Examples of the story‐board designs
49. The problem of VAD remains :
GoldenRice is a potentially
significant contribution to alleviation.
It is now on its way towards registration
50. GR2 GR1
Wild-Type
For additional information, see www.goldenrice.org
51. Philippines
generally high VAD, (and increasing) especially Visayas & W Mindanao
(figures are % of that population with VAD)
Children, 6 Months to 5 Years Pregnant Women Lactating Women
Source: Philippine Government Statistics 1998, 2003