In-Vitro Pollination and Fertilization
The document discusses in-vitro pollination and fertilization techniques. It begins with a brief history, noting its development in 1902 and use to produce hybrids between incompatible species. It then describes barriers to pollination and fertilization that can be overcome through in-vitro methods. Several techniques are outlined, including ovule, ovary, and stigma cultures. Requirements for successful in-vitro fertilization include viable gametes and proper culture conditions. The document concludes by discussing applications in plant breeding like overcoming self-incompatibility and producing stress-tolerant hybrids.
Anther culture is a technique where anthers are excised from flower buds and cultured to produce haploid plants. The first report of haploid tissue from anther culture was in 1964-1966 in Datura pollen grains. Over 250 species have been produced through anther culture, most commonly in families like Solanaceae, Cruciferae, and Poaceae. Haploid plants are useful for identifying recessive traits, eliminating lethal genes, and producing homozygous diploid plants more quickly. There are several pathways that microspores can follow during anther culture, such as symmetric or asymmetric division, to produce haploid plants. Successful anther culture requires optimizing various factors like donor plant genotype, anther
Meristem tip culture for the production of the virus free plantsArjun Rayamajhi
This presentation gives general idea on the meristem tip culture for the production of the virus free plants. The principles, methods and procedures of the meristem tip culture included. General idea on different in vitro culture techniques for virus elimination meristem tip culture viz. thermotherapy, cryotherapy,chemotherapy and electrotherapy are provided.
This document discusses different types of plant organ culture, including root, shoot apical meristem, leaf, flower, and ovule cultures. Root culture involves culturing excised radical tips of aseptically germinated seeds. Shoot apical meristem culture involves culturing the shoot tip comprising the meristem and developing leaves. Flower culture involves culturing excised floral buds to produce full blooms. Ovule culture involves culturing isolated ovules to facilitate fertilization and embryo development. Organ cultures have various applications including studying organ development, production of secondary metabolites, and generating virus-free plants.
Invitro culture of unpollinated ovaries and ovules represents an alternative for the production of haploid plant
First successful report on the induction of gynogenic haploid was in barley by San Noeum in 1976
Haploid plants are obtained from ovary and ovule culture of rice, wheat, maize, sunflower, tobacco, poplar, mulberry etc
Whites or MS or N6 inorganic salt medium supplement with growth substances are used
The document discusses organogenesis, which is the development of adventitious organs or primordial from undifferentiated plant cell mass through differentiation. It describes the process, including dedifferentiation and redifferentiation stages. There are two types of organogenesis - direct organogenesis which does not involve callus formation, and indirect organogenesis which involves callus formation. Organogenesis is used in plant tissue culture to regenerate plants through shoot or root cultures and is influenced by factors like explant source and size, plant growth regulators, and culture conditions. It has commercial applications in micropropagation of plants.
This document discusses anther and pollen culture techniques. It provides a brief history of the development of these techniques from the 1950s onward. It then describes the process of anther culture, where anthers are cultured in nutrient medium to produce haploid callus or embryos. Pollen or microspore culture involves isolating pollen grains from anthers and culturing them. The goal is to produce haploid embryos or callus that can develop into haploid plantlets. Key factors that influence success include the genotype, microspore stage, culture medium, temperature, and physiological status of the donor plant. Anther culture has applications in mutation studies, plant breeding, and secondary metabolite production.
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).
In-Vitro Pollination and Fertilization
The document discusses in-vitro pollination and fertilization techniques. It begins with a brief history, noting its development in 1902 and use to produce hybrids between incompatible species. It then describes barriers to pollination and fertilization that can be overcome through in-vitro methods. Several techniques are outlined, including ovule, ovary, and stigma cultures. Requirements for successful in-vitro fertilization include viable gametes and proper culture conditions. The document concludes by discussing applications in plant breeding like overcoming self-incompatibility and producing stress-tolerant hybrids.
Anther culture is a technique where anthers are excised from flower buds and cultured to produce haploid plants. The first report of haploid tissue from anther culture was in 1964-1966 in Datura pollen grains. Over 250 species have been produced through anther culture, most commonly in families like Solanaceae, Cruciferae, and Poaceae. Haploid plants are useful for identifying recessive traits, eliminating lethal genes, and producing homozygous diploid plants more quickly. There are several pathways that microspores can follow during anther culture, such as symmetric or asymmetric division, to produce haploid plants. Successful anther culture requires optimizing various factors like donor plant genotype, anther
Meristem tip culture for the production of the virus free plantsArjun Rayamajhi
This presentation gives general idea on the meristem tip culture for the production of the virus free plants. The principles, methods and procedures of the meristem tip culture included. General idea on different in vitro culture techniques for virus elimination meristem tip culture viz. thermotherapy, cryotherapy,chemotherapy and electrotherapy are provided.
This document discusses different types of plant organ culture, including root, shoot apical meristem, leaf, flower, and ovule cultures. Root culture involves culturing excised radical tips of aseptically germinated seeds. Shoot apical meristem culture involves culturing the shoot tip comprising the meristem and developing leaves. Flower culture involves culturing excised floral buds to produce full blooms. Ovule culture involves culturing isolated ovules to facilitate fertilization and embryo development. Organ cultures have various applications including studying organ development, production of secondary metabolites, and generating virus-free plants.
Invitro culture of unpollinated ovaries and ovules represents an alternative for the production of haploid plant
First successful report on the induction of gynogenic haploid was in barley by San Noeum in 1976
Haploid plants are obtained from ovary and ovule culture of rice, wheat, maize, sunflower, tobacco, poplar, mulberry etc
Whites or MS or N6 inorganic salt medium supplement with growth substances are used
The document discusses organogenesis, which is the development of adventitious organs or primordial from undifferentiated plant cell mass through differentiation. It describes the process, including dedifferentiation and redifferentiation stages. There are two types of organogenesis - direct organogenesis which does not involve callus formation, and indirect organogenesis which involves callus formation. Organogenesis is used in plant tissue culture to regenerate plants through shoot or root cultures and is influenced by factors like explant source and size, plant growth regulators, and culture conditions. It has commercial applications in micropropagation of plants.
This document discusses anther and pollen culture techniques. It provides a brief history of the development of these techniques from the 1950s onward. It then describes the process of anther culture, where anthers are cultured in nutrient medium to produce haploid callus or embryos. Pollen or microspore culture involves isolating pollen grains from anthers and culturing them. The goal is to produce haploid embryos or callus that can develop into haploid plantlets. Key factors that influence success include the genotype, microspore stage, culture medium, temperature, and physiological status of the donor plant. Anther culture has applications in mutation studies, plant breeding, and secondary metabolite production.
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).
Synthetic seeds are encapsulated somatic embryos or shoot buds that can be used for planting like traditional seeds. They allow for clonal propagation of plants that are difficult to reproduce through traditional seeds, including some fruit crops. The production of synthetic seeds involves inducing somatic embryogenesis in callus cultures, maturing the embryos, and encapsulating them in a protective gel before planting. This allows genetic material to be stored and dispersed while avoiding issues with seed-borne diseases, low seed viability, and difficulties reproducing species that lack traditional seeds.
This document summarizes research on somatic embryogenesis in rice. It describes the process of somatic embryogenesis, including the stages of embryogenesis and factors that affect it. The methodology section outlines the materials and methods used, including collecting rice seeds as explants, sterilizing them, and culturing them on callus induction and embryo germination media with different concentrations of plant growth regulators like 2,4-D, BAP and NAA. The goal is to develop an efficient system for somatic embryogenesis and plant regeneration in rice.
Plant tissue culture is a technique used to propagate plants asexually under controlled laboratory conditions. It involves culturing plant cells, tissues, or organs on artificial nutrient media. The key principles are totipotency, where plant cells can regenerate into whole plants, and plasticity, where plants can alter their growth to suit their environment. There are various types of plant tissue culture including embryo, seed, meristem, cell, protoplast, callus, pollen, and organ culture. Important applications include producing virus-free plants through meristem culture and generating novel hybrids through protoplast fusion.
Introduction to organ culture in ptc and root cultureCollege
This presentation gives details about the organ culture in plant tissue culture and its basic applications, also this provide an detailed information about the technique of root culture and gives small view about its appilications.
Introduction
Reason for cryopreservation
Selection of part of plant for cryopreservation
Technique of cryopreservation
Application
Limitation
Conclusion
This practical aims to induce callus formation from different plant explants. Students will take explants like leaf, stem, and root segments from selected horticultural crops and culture them on nutrient media supplemented with plant growth regulators, like auxins and cytokinins, to induce callus tissue. The success of callus induction will depend on the explant type and plant species as well as the plant growth regulator concentrations used in the culture medium.
Micropropagation is the process of rapidly multiplying stock plant materials using modern tissue culture methods. It involves 5 main stages: selection of stock plants, initiation and establishment of culture, multiplication of shoots, rooting of shoots, and establishment of plantlets. The two main approaches are multiplication through axillary buds/apical shoots and adventitious shoots. Organogenesis and somatic embryogenesis are also used, where organs or embryos are formed directly or indirectly from explants. Micropropagation is useful for cloning noble plants and providing sufficient plantlets from stock plants that do not produce many seeds or vegetatively propagate well.
Tissue culture is a technique where cells, tissues or whole plants are grown in a sterile nutrient culture medium under controlled conditions. It allows for rapid vegetative propagation of plants. Key steps include sterilizing equipment and explants, preparing nutrient medium, subculturing to promote growth, and rooting and hardening plantlets. Tissue culture has many applications like mass multiplication of crops and plants, eliminating diseases, and genetic modification. It is used commercially for propagating crops but contamination and rooting difficulties can be issues.
Plant tissue culture involves cultivating plant cells, tissues, or organs in an artificial nutrient medium under sterile conditions. It relies on the totipotency and plasticity of plant cells. Explant material is surface sterilized and transferred to nutrient medium containing sugars, salts, vitamins, and plant growth regulators. Under the right conditions, callus tissue will form and develop into shoots and roots, allowing for the regeneration of whole plants through micropropagation. Plant tissue culture has many advantages, including the ability to rapidly produce large numbers of genetically identical plants, recover healthy plants from diseased stock, and develop new varieties through genetic engineering techniques.
Somatic embryogenesis, in plant tissue culture 2KAUSHAL SAHU
Introduction
Types of somatic embryogenesis
Developmental stages
Factors affecting somatic embryogenesis
Importance
Conclusions
References
The process of regeneration of embryos from somatic cells, tissue or organs is regarded as somatic or asexual embryogenesis.
opposite of zygotic or sexual embryogenesis.
Embryo-like structures which can develop into whole plants in a way that is similar to zygotic embryos are formed from somatic cells.
OVARY CULTURE:-
"the in-vitro culturing of ovaries in an aseptic condition from the pollinated or un-pollinated flowers, in an appropriate nutrient medium and under optimal conditions." And
OVULE CULTURE:-
"Ovule culture is an experimental system by which ovules are aseptically isolated from the ovary and are grown aseptically on chemically defined nutrient medium under controlled conditions."
This document discusses using in vitro mutagenesis techniques to induce desirable mutations in flower crops. It describes mutagenesis as a process that changes genetic information, resulting in mutations. Both physical (radiation) and chemical (EMS, DES) mutagens are used. The document presents case studies on inducing mutations in tuberose and rose plants using gamma rays and chemical mutagens. Desirable variations were observed, such as changes in flower shape, color, and stem length. Radiation doses of 40 Gy and 55 Gy were found to successfully induce novel variants in rose. The objective is to develop improved varieties of ornamental plants using these mutation techniques.
Callus is an unorganized mass of undifferentiated cells that can be cultured in vitro. It is produced when plant explants are cultured on medium containing auxin and cytokinin hormones under sterile conditions. Callus tissue lacks differentiation and is unable to perform photosynthesis. It can be maintained indefinitely and used for plant regeneration through processes like organogenesis and somatic embryogenesis. Successful callus culture requires aseptic preparation of explants, a nutrient medium with proper hormone balance, and controlled physical conditions for incubation.
Regeneration of plants and application of plant tissue culture SuruchiDahiya
Plant tissue culture is a collection of techniques used to grow plant cells, tissues, or organs in sterile conditions. It relies on the fact that many plant cells can regenerate a whole plant. There are several types of tissue culture including callus culture, organ culture, and suspension culture. Plant tissue culture has many applications including micropropagation, production of pharmaceuticals, and genetic engineering of plants. It is a valuable tool for producing disease-free plants and increasing crop yields.
The document discusses plant tissue culture media. It describes media as a substance that supports and provides energy and nutrients for developing plant explants. The main types of media are solid and liquid, and their composition varies depending on the explant species. Common media include Murashige and Skoog, White's Medium, Gamborg Medium, and Nitsch's Medium. Media contains inorganic supplements like macronutrients and micronutrients as well as organic supplements such as vitamins, amino acids, and energy sources. Other constituents include growth regulators, solidifying agents, and pH buffers. Media is prepared by making stock solutions of dry and wet ingredients, adding a gelling agent, and sterilizing the final mixture.
Plant tissue culture is the process of growing plant cells, tissues or organs in sterile conditions on a nutrient medium. It has many applications like germplasm preservation of endangered plants, genetic improvement of crops, and production of secondary metabolites. The basic steps include selection of explant, initiation of culture on growth media, multiplication through cell division, and rooting and transfer to soil. Technologies such as micropropagation, somatic hybridization, and cryopreservation have been commercialized for mass propagation of crops. However, issues remain regarding genetic stability, selection of fused products, and regeneration efficiency.
Hardening, packaging & transport of micropropagules and construction of p...AjaykumarKarna
1. The document discusses various techniques for hardening, packaging, transporting, and propagating tissue cultured plants, including micropropagules.
2. It describes hardening processes, various packaging materials and methods, and considerations for transporting tissue cultured plants by cargo.
3. Propagation structures that are discussed include greenhouses, hot beds, cold frames, lath houses, propagation frames, net houses, bottom heat boxes, and mist propagation units - each with specific purposes and construction details provided.
This document summarizes ovary and ovule culture techniques. Ovary culture involves growing unpollinated flower ovaries in nutrient medium to produce haploid plants through gynogenesis. Ovule culture involves isolating ovules from ovaries and culturing them separately. The techniques allow studying early embryo development, effects of plant hormones, and fruit physiology. Protocols involve surface sterilizing and culturing ovaries or isolating and culturing ovules on solid or liquid nutrient medium. The cultures are incubated at 25°C and can be grown in light or dark conditions. Ovary and ovule culture have applications in plant breeding and research.
Unit 3.0 introduction and history of plant tissue cultureDr. Mafatlal Kher
Plant tissue culture is the process of growing plant cells, tissues or organs in an artificial nutrient medium under sterile conditions. The document discusses the history and development of plant tissue culture techniques. It notes that plant tissue culture is founded on cell theory proposed by Schleiden and Schwann in 1838-1839. Gottlieb Haberlandt is considered the father of plant tissue culture for his pioneering experiments in 1902, though his experiments failed due to inappropriate plant material and nutrient medium. Improved nutrient solutions like Knop's solution, White's medium and Murashige and Skoog medium enabled indefinite growth and multiplication of plant tissues in culture.
1) Shoot tip culture involves culturing the terminal portion of a shoot tip, comprising the meristem and developing leaves and stem tissue.
2) It is used to produce virus-free plants by removing viruses that cannot move between cells, for micropropagation, and to store plant genetic resources.
3) The protocol involves surface sterilizing and culturing small shoot tip explants through stages of culture establishment, shoot proliferation, and root regeneration. Factors like explant size and physiological condition affect the process.
The document summarizes the operationalization of a tissue culture lab and farm in Chauntra, Himachal Pradesh through a public-private partnership between the DRDA Mandi, CSIR-IHBT Palampur, and GreenTech Agri-Sector Pvt. Ltd. The partnership aims to commercially produce economically important plants using advanced tissue culture techniques to benefit farmers. GreenTech will manage the operational aspects of the lab and farm with support from DRDA for infrastructure development and technology transfer from CSIR-IHBT.
The document provides information about plant tissue culture including the history, basic procedures, requirements for establishing a tissue culture laboratory, common terms used, and advantages and disadvantages. It describes the key steps of taking an explant from a mother plant, culturing it in a sterile nutrient medium, and multiplying the plantlets. The goal of tissue culture is to produce many genetically identical copies of desirable plants in a controlled, contaminant-free environment.
Synthetic seeds are encapsulated somatic embryos or shoot buds that can be used for planting like traditional seeds. They allow for clonal propagation of plants that are difficult to reproduce through traditional seeds, including some fruit crops. The production of synthetic seeds involves inducing somatic embryogenesis in callus cultures, maturing the embryos, and encapsulating them in a protective gel before planting. This allows genetic material to be stored and dispersed while avoiding issues with seed-borne diseases, low seed viability, and difficulties reproducing species that lack traditional seeds.
This document summarizes research on somatic embryogenesis in rice. It describes the process of somatic embryogenesis, including the stages of embryogenesis and factors that affect it. The methodology section outlines the materials and methods used, including collecting rice seeds as explants, sterilizing them, and culturing them on callus induction and embryo germination media with different concentrations of plant growth regulators like 2,4-D, BAP and NAA. The goal is to develop an efficient system for somatic embryogenesis and plant regeneration in rice.
Plant tissue culture is a technique used to propagate plants asexually under controlled laboratory conditions. It involves culturing plant cells, tissues, or organs on artificial nutrient media. The key principles are totipotency, where plant cells can regenerate into whole plants, and plasticity, where plants can alter their growth to suit their environment. There are various types of plant tissue culture including embryo, seed, meristem, cell, protoplast, callus, pollen, and organ culture. Important applications include producing virus-free plants through meristem culture and generating novel hybrids through protoplast fusion.
Introduction to organ culture in ptc and root cultureCollege
This presentation gives details about the organ culture in plant tissue culture and its basic applications, also this provide an detailed information about the technique of root culture and gives small view about its appilications.
Introduction
Reason for cryopreservation
Selection of part of plant for cryopreservation
Technique of cryopreservation
Application
Limitation
Conclusion
This practical aims to induce callus formation from different plant explants. Students will take explants like leaf, stem, and root segments from selected horticultural crops and culture them on nutrient media supplemented with plant growth regulators, like auxins and cytokinins, to induce callus tissue. The success of callus induction will depend on the explant type and plant species as well as the plant growth regulator concentrations used in the culture medium.
Micropropagation is the process of rapidly multiplying stock plant materials using modern tissue culture methods. It involves 5 main stages: selection of stock plants, initiation and establishment of culture, multiplication of shoots, rooting of shoots, and establishment of plantlets. The two main approaches are multiplication through axillary buds/apical shoots and adventitious shoots. Organogenesis and somatic embryogenesis are also used, where organs or embryos are formed directly or indirectly from explants. Micropropagation is useful for cloning noble plants and providing sufficient plantlets from stock plants that do not produce many seeds or vegetatively propagate well.
Tissue culture is a technique where cells, tissues or whole plants are grown in a sterile nutrient culture medium under controlled conditions. It allows for rapid vegetative propagation of plants. Key steps include sterilizing equipment and explants, preparing nutrient medium, subculturing to promote growth, and rooting and hardening plantlets. Tissue culture has many applications like mass multiplication of crops and plants, eliminating diseases, and genetic modification. It is used commercially for propagating crops but contamination and rooting difficulties can be issues.
Plant tissue culture involves cultivating plant cells, tissues, or organs in an artificial nutrient medium under sterile conditions. It relies on the totipotency and plasticity of plant cells. Explant material is surface sterilized and transferred to nutrient medium containing sugars, salts, vitamins, and plant growth regulators. Under the right conditions, callus tissue will form and develop into shoots and roots, allowing for the regeneration of whole plants through micropropagation. Plant tissue culture has many advantages, including the ability to rapidly produce large numbers of genetically identical plants, recover healthy plants from diseased stock, and develop new varieties through genetic engineering techniques.
Somatic embryogenesis, in plant tissue culture 2KAUSHAL SAHU
Introduction
Types of somatic embryogenesis
Developmental stages
Factors affecting somatic embryogenesis
Importance
Conclusions
References
The process of regeneration of embryos from somatic cells, tissue or organs is regarded as somatic or asexual embryogenesis.
opposite of zygotic or sexual embryogenesis.
Embryo-like structures which can develop into whole plants in a way that is similar to zygotic embryos are formed from somatic cells.
OVARY CULTURE:-
"the in-vitro culturing of ovaries in an aseptic condition from the pollinated or un-pollinated flowers, in an appropriate nutrient medium and under optimal conditions." And
OVULE CULTURE:-
"Ovule culture is an experimental system by which ovules are aseptically isolated from the ovary and are grown aseptically on chemically defined nutrient medium under controlled conditions."
This document discusses using in vitro mutagenesis techniques to induce desirable mutations in flower crops. It describes mutagenesis as a process that changes genetic information, resulting in mutations. Both physical (radiation) and chemical (EMS, DES) mutagens are used. The document presents case studies on inducing mutations in tuberose and rose plants using gamma rays and chemical mutagens. Desirable variations were observed, such as changes in flower shape, color, and stem length. Radiation doses of 40 Gy and 55 Gy were found to successfully induce novel variants in rose. The objective is to develop improved varieties of ornamental plants using these mutation techniques.
Callus is an unorganized mass of undifferentiated cells that can be cultured in vitro. It is produced when plant explants are cultured on medium containing auxin and cytokinin hormones under sterile conditions. Callus tissue lacks differentiation and is unable to perform photosynthesis. It can be maintained indefinitely and used for plant regeneration through processes like organogenesis and somatic embryogenesis. Successful callus culture requires aseptic preparation of explants, a nutrient medium with proper hormone balance, and controlled physical conditions for incubation.
Regeneration of plants and application of plant tissue culture SuruchiDahiya
Plant tissue culture is a collection of techniques used to grow plant cells, tissues, or organs in sterile conditions. It relies on the fact that many plant cells can regenerate a whole plant. There are several types of tissue culture including callus culture, organ culture, and suspension culture. Plant tissue culture has many applications including micropropagation, production of pharmaceuticals, and genetic engineering of plants. It is a valuable tool for producing disease-free plants and increasing crop yields.
The document discusses plant tissue culture media. It describes media as a substance that supports and provides energy and nutrients for developing plant explants. The main types of media are solid and liquid, and their composition varies depending on the explant species. Common media include Murashige and Skoog, White's Medium, Gamborg Medium, and Nitsch's Medium. Media contains inorganic supplements like macronutrients and micronutrients as well as organic supplements such as vitamins, amino acids, and energy sources. Other constituents include growth regulators, solidifying agents, and pH buffers. Media is prepared by making stock solutions of dry and wet ingredients, adding a gelling agent, and sterilizing the final mixture.
Plant tissue culture is the process of growing plant cells, tissues or organs in sterile conditions on a nutrient medium. It has many applications like germplasm preservation of endangered plants, genetic improvement of crops, and production of secondary metabolites. The basic steps include selection of explant, initiation of culture on growth media, multiplication through cell division, and rooting and transfer to soil. Technologies such as micropropagation, somatic hybridization, and cryopreservation have been commercialized for mass propagation of crops. However, issues remain regarding genetic stability, selection of fused products, and regeneration efficiency.
Hardening, packaging & transport of micropropagules and construction of p...AjaykumarKarna
1. The document discusses various techniques for hardening, packaging, transporting, and propagating tissue cultured plants, including micropropagules.
2. It describes hardening processes, various packaging materials and methods, and considerations for transporting tissue cultured plants by cargo.
3. Propagation structures that are discussed include greenhouses, hot beds, cold frames, lath houses, propagation frames, net houses, bottom heat boxes, and mist propagation units - each with specific purposes and construction details provided.
This document summarizes ovary and ovule culture techniques. Ovary culture involves growing unpollinated flower ovaries in nutrient medium to produce haploid plants through gynogenesis. Ovule culture involves isolating ovules from ovaries and culturing them separately. The techniques allow studying early embryo development, effects of plant hormones, and fruit physiology. Protocols involve surface sterilizing and culturing ovaries or isolating and culturing ovules on solid or liquid nutrient medium. The cultures are incubated at 25°C and can be grown in light or dark conditions. Ovary and ovule culture have applications in plant breeding and research.
Unit 3.0 introduction and history of plant tissue cultureDr. Mafatlal Kher
Plant tissue culture is the process of growing plant cells, tissues or organs in an artificial nutrient medium under sterile conditions. The document discusses the history and development of plant tissue culture techniques. It notes that plant tissue culture is founded on cell theory proposed by Schleiden and Schwann in 1838-1839. Gottlieb Haberlandt is considered the father of plant tissue culture for his pioneering experiments in 1902, though his experiments failed due to inappropriate plant material and nutrient medium. Improved nutrient solutions like Knop's solution, White's medium and Murashige and Skoog medium enabled indefinite growth and multiplication of plant tissues in culture.
1) Shoot tip culture involves culturing the terminal portion of a shoot tip, comprising the meristem and developing leaves and stem tissue.
2) It is used to produce virus-free plants by removing viruses that cannot move between cells, for micropropagation, and to store plant genetic resources.
3) The protocol involves surface sterilizing and culturing small shoot tip explants through stages of culture establishment, shoot proliferation, and root regeneration. Factors like explant size and physiological condition affect the process.
The document summarizes the operationalization of a tissue culture lab and farm in Chauntra, Himachal Pradesh through a public-private partnership between the DRDA Mandi, CSIR-IHBT Palampur, and GreenTech Agri-Sector Pvt. Ltd. The partnership aims to commercially produce economically important plants using advanced tissue culture techniques to benefit farmers. GreenTech will manage the operational aspects of the lab and farm with support from DRDA for infrastructure development and technology transfer from CSIR-IHBT.
The document provides information about plant tissue culture including the history, basic procedures, requirements for establishing a tissue culture laboratory, common terms used, and advantages and disadvantages. It describes the key steps of taking an explant from a mother plant, culturing it in a sterile nutrient medium, and multiplying the plantlets. The goal of tissue culture is to produce many genetically identical copies of desirable plants in a controlled, contaminant-free environment.
This document contains protocols for various plant tissue culture techniques. It discusses the introduction to plant tissue culture, sterilization techniques used, and then outlines 8 specific protocols: 1) tissue culture media preparation, 2) explant preparation and surface sterilization, 3) embryo culture, 4) culture of anther for haploid production, 5) meristem culture, 6) meristem tip culture for virus-free plants, 7) induction of somatic embryogenesis, and 8) protoplast isolation, culture, and regeneration. The goal of these protocols is to describe the principles and procedures of different plant tissue culture methods.
Plant tissue culture involves growing plant cells, tissues, or organs in sterile conditions on a nutrient culture medium. Key aspects of plant tissue culture include selecting and sterilizing explant tissue, transferring it to a solid culture medium containing hormones, and periodically moving the growing tissue to new media for multiplication. The growing plantlets are eventually transferred to pots and grown in a greenhouse. Plant tissue culture allows for conservation of endangered species, production of virus-free plants, hybrid regeneration, and faster plant growth.
The presentation covered the basic steps of plant tissue culture, including setting up a tissue culture lab with areas for media preparation, aseptic transfer, culture, analysis, and acclimatization. It discussed media components and preparation, sterilization techniques like filtration, radiation, and chemicals, and sterilizing plant tissues using chemicals like sodium hypochlorite. Proper cleaning of glassware and sterilization of equipment and materials are essential for maintaining aseptic conditions.
The document discusses the requirements for setting up a low-cost plant tissue culture laboratory. It outlines the necessary structural elements like proper roofing, drainage, and ventilation. Key laboratory rooms include washing, sterilization, media preparation, inoculation, and incubation rooms. Essential equipment includes an autoclave, laminar airflow cabinet, incubators, water purification system, balances, pH meter, and glassware. The total estimated cost for establishing such a laboratory is around Rs. 150,000 to 200,000. Proper facilities for hand washing, fire safety, and waste disposal should also be provided.
The document discusses the essential components of plant tissue culture media, including macro and micro nutrients, organic nutrients like vitamins and carbohydrates, and plant growth regulators. It explains that the success of tissue culture depends on using the right type of culture media, which must contain nutrients, carbon sources, and other components to support in vitro plant growth. The various roles and forms of important media components like nitrogen, calcium, iron, and cytokinins are also outlined.
This document provides an overview of plant tissue culture techniques. It discusses that plant tissue culture involves growing plant cells, tissues or organs in a sterile environment with nutrient media. The techniques rely on two principles - totipotency, the ability of plant cells to regenerate into a whole plant, and plasticity, the ability of plants to alter their growth in response to their environment. Explants from various plant tissues are sterilized and placed on culture media, which are composed of inorganic salts, organic nutrients and plant hormones. The culture media, explant source, and plant species can affect regeneration efficiency. Applications of plant tissue culture include commercial plant production, conservation of rare species, screening for desirable traits like herbicide resistance, and
Plant tissue culture is the process of growing plant cells, tissues or organs in vitro under sterile conditions on a nutrient medium. The father of plant tissue culture was Haberlandt who first conceived of culturing plant cells aseptically in 1902. There are two main types of growth in tissue culture - organized growth where structure is preserved, and unorganized growth like callus or cell suspension cultures which lack structure. Key steps in establishing tissue cultures include selecting an explant, surface sterilization, and culturing on solid or liquid media. Plant growth regulators like auxins and cytokinins are important for directing growth. Tissue cultures are used to produce valuable secondary metabolites like taxol through optimization of culture conditions and addition of elic
This document provides an overview of plant tissue culture. It defines tissue culture as the in vitro cultivation of plant cells or tissues under aseptic conditions on a nutrient medium. The history and key figures in the development of plant tissue culture are discussed. Details are provided on nutrient requirements, preparation and sterilization of culture media, basic laboratory requirements, establishment of cultures from explants, and types of growth and cultures. The advantages and applications of plant tissue culture are also summarized.
basic principles and protocol in plant tissue cultureAlia Najiha
Plant tissue culture involves growing plant cells, tissues, or organs in an artificial nutrient medium under sterile conditions. It allows for the rapid production of exact genetic copies of plants and is used to regenerate whole plants from genetically modified plant cells. The key requirements for successful tissue culture are appropriate explant tissue, a suitable growth medium, aseptic conditions to prevent microbial contamination, growth regulators, and frequent subculturing to provide nutrients and remove waste. The basic steps are selection of explant tissue from a healthy mother plant, sterilization of the tissue, establishment of the explant in a culture medium, multiplication through callus formation and shoot development, and root formation through manipulation of growth regulators. Benefits include providing pathogen-free plants
The document discusses several human diseases including their causes, symptoms, transmission, prevention and control. It covers viral diseases like influenza, jaundice, rabies and AIDS. It also discusses bacterial diseases such as cholera, diarrhea and typhoid. Additionally, it summarizes protozoan diseases including malaria and parasitic diseases like tuberculosis. The document provides detailed information on the definition, characteristics, treatment and prevention of major infectious diseases impacting humans.
This document discusses media used for plant tissue culture. It provides details on the components and purpose of various standard media including Murashige and Skoog medium, Gamborg's B5 medium, Nitsch medium, and others. It also discusses the composition and use of specialized media for banana, orchid, and woody plant tissue culture. The key components of media include minerals, vitamins, carbon sources, and plant growth regulators to provide nutrients and regulate growth for cultured plant cells and tissues.
PPT on Tissue Culture Class 10 CBSE Text Book NCERT.One Time Forever
This is a PPT Based on Class 10 Chapter How Do Organisms Reproduce, on a Small Topic of it That is Tissue Culture with easy and detailed explanation of each topic of tissue culture along with some pictures and some examples. Hopefully it Would Be Helpful To You. Thank You.
This document summarizes a student group's plant tissue culture project on banana propagation. It includes:
1) The group members and their lecturer. The objective is to understand plant tissue culture procedures.
2) Banana suckers are commonly used as explants in tissue culture. The document describes sterilization and preparation of MS media for culturing banana shoot tips.
3) The results showed both contaminated and uncontaminated cultures. Factors affecting contamination and growth are discussed. Suggestions for improvement include using meristematic shoot tips and adding antioxidants to the media.
The document discusses secondary metabolite production using plant tissue culture techniques. It describes how secondary metabolites are organic compounds not directly involved in growth that play roles in plant defense. Plant tissue culture and genetic engineering methods can be used to control secondary metabolite production, including manipulating the environment, growth conditions, precursor addition, and transforming plant cells with genes from bacteria like Agrobacterium that influence metabolite pathways. Overall, the document provides an overview of how secondary metabolites are classified and various biotechnological approaches for enhancing their production in plant cell cultures.
This seminar presentation summarizes anther culture and haploid plant regeneration. Anther culture involves culturing microspores or anthers to produce haploid plants. It has been successful in over 250 plant species. There are different pathways that microspores can take to develop into haploid plants, including equal or unequal division. Pretreatment of anthers and culture conditions like medium, temperature and light exposure can influence success rates. Anther culture is useful for producing haploids but requires skill and has risks of chimera or callus formation.
This document is a biology project on immunity submitted by Rahul Tripathi of class XII to his teacher Mr. Vikas Dixit. It contains an introduction, table of contents, chapters on the immune system including the fluid system, granulocytes, agranulocytes, lymphocytes, defense mechanisms, types of immunity like innate and adaptive immunity, and a conclusion thanking the teacher. The project aims to explain the key concepts of the immune system through chapters on its various components and functions to build understanding of how the body resists diseases.
Plant tissue culture is the in vitro cultivation of plant cells, tissues, or organs on defined nutrient media under sterile conditions. It allows plants to be grown free from diseases and produced artificially in large quantities. The document discusses the history, basic requirements, types (callus culture, suspension culture), applications (clonal propagation, secondary metabolite production, genetic variability), and edible vaccines of plant tissue culture.
Mass multiplication procedure for tissue culture and PTC requirementDr. Deepak Sharma
This presentation include basic Micropropagation protocol: Application and advantages of mass multiplication. Beside this the requirement of tissue culture are there (Nutrient, gelling agent, energy source, vitamins and PGRs) are also included.
The document discusses various tissue culture techniques used in plant breeding including: clonal propagation of disease-free genetic stocks through tissue culture; freeze preservation of germplasm; embryo, ovule, and anther culture techniques to produce haploid plants; and the induction of genetic variability through cell cultures. It provides details on the basic procedures of plant tissue culture including establishment of aseptic culture from explants, proliferation of callus cells on nutrient media, rooting, and acclimatization of regenerated plantlets. The roles of growth hormones, nutrient media composition, and factors affecting culture efficiency are also summarized.
Application of plant tissue culture/ micro-propagationSushil Nyaupane
Tissue culture is the process of growing cells or tissues in sterile conditions. It allows for rapid cloning of plant materials. Plant tissue culture involves excising plant parts and growing them on nutrient media. This allows for mass multiplication of plant materials irrespective of season. Some key developments include Haberlandt's proposal of plant cell culture in 1902, and Murashige and Skoog's nutrient medium in 1962. Micropropagation is now used for conservation of rare species, producing disease-free plants, mutation breeding, and more. The future of this technique remains promising.
Plant tissue culture is the process of culturing plant cells, tissues or organs in a nutrient medium under sterile conditions. It has many applications in agriculture including producing rare hybrids, disease-free plants through embryo culture, and micropropagation for vegetative propagation. The key steps involve selecting an explant, sterilizing it, inoculating it in a nutrient medium, initiating callus growth, subculturing, regenerating plantlets, and hardening them for transfer. Plant tissue culture offers advantages like rapid multiplication of plants with optimal traits but has disadvantages like lack of genetic variation. It has potential to commercialize important crops and ensure food security through breeding improved varieties.
I have discussed Applications of Plant Tissue Culture under the following subheadings,
1. Micro Propagation
2. Clonal Propagation
3. Production of Genetically Variable Plants
4. Production of Virus Free Plants
5. Plant Breeding
6. Production of Useful Biochemicals
7. Preservation of Plant Genetic Resources
8. Importance of Tissue Culture in Biotechnology
This document provides an overview of plant tissue culture methods and applications. It discusses the basic concepts of plant tissue culture, including plasticity and totipotency. The stages of micropropagation are outlined as initiation, multiplication, rooting, and transfer to soil. Micropropagation involves regeneration and multiplication of plants from explants like axillary buds and shoot tips. The document also discusses meristem culture, shoot tip culture, growth media, factors affecting tissue culture, applications like disease elimination and germplasm conservation, somaclonal variation, and cryopreservation.
Plant tissue culture has been widely employed in area of agriculture, horticulture, forestry and plant breeding. It is an applied biotechnology used for mass propagation, virus elimination, secondary metabolite production and in vitro cloning of plants. Recently, plant tissue culture has been used for the conservation of endangered plant species through short and medium term conservation also known as slow growth and cryopreservation also known as long term conservation. These methods had been effectively used to conserve plant species with recalcitrant seeds or dormant seeds and showed greater advantage over the conventional methods of conservation. At present plant cell culture has made great advances. Possibly the most significant role that plant cell culture has to play in the future will be in its association with transgenic plants. The ability to accelerate the conventional multiplication rate can be of great benefit to many crops countries where a disease or some climatic disaster wipes out crops. Mr. Rohan R. Vakhariya | Rutuja R. Shah "Over Review on Plant Tissue Culture" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-4 | Issue-1 , December 2019, URL: https://www.ijtsrd.com/papers/ijtsrd29619.pdfPaper URL: https://www.ijtsrd.com/pharmacy/other/29619/over-review-on-plant-tissue-culture/mr-rohan-r-vakhariya
Tissue culture is a process that clones plants through micropropagation. It involves culturing plant tissues in sterile conditions with specific nutrients and hormones. There are four main stages - initiation, multiplication, rooting, and acclimatization. The multiplication stage uses cytokinins to induce shoot growth from explants like leaves or stems. Rooting uses auxins to induce root formation from shoots. The process allows for mass production of genetically identical plants independent of seasons.
Plant tissue culture is a collection of techniques used to maintain or grow plant cells, tissues, and organs under sterile conditions. The history of plant tissue culture began in the 1830s with theories of cell totipotency. Significant developments included the discovery of plant growth regulators in the 1920s-1940s and the development of plant cell differentiation and somatic embryogenesis in the 1950s-1960s. There are several types of plant tissue culture including shoot culture, callus culture, embryo culture, and meristem culture. Applications include germplasm conservation, large-scale production, disease eradication, genetic engineering, and more. The advantages are rapid propagation, disease-free plants, year-round growth, and conservation of endangered
Plant tissue culture is the process of growing plant cells, tissues or organs in sterile conditions on a nutrient medium. It has many applications like germplasm preservation of endangered plants, producing disease-free plants through micropropagation, and creating novel hybrids through protoplast fusion and somatic hybridization. However, issues remain like genetic instability of hybrids and lack of efficient selection methods. Overall, tissue culture is a valuable biotechnology tool with potential for crop improvement and conservation efforts.
Secondary metabolites are organic compounds produced by plant metabolism that are not essential for growth or reproduction but provide other benefits. They often function in plant defense against herbivores and pathogens. There are several types of plant tissue cultures used to study secondary metabolism, including organized cultures of tissues, disorganized callus cultures, hairy root cultures, and immobilized cell cultures where cells are confined within a matrix.
This document discusses micropropagation as a method of clonally propagating plants. It begins by explaining traditional clonal propagation methods and their limitations. It then describes the benefits of micropropagation, which allows for rapid multiplication of plants using small explant tissues in sterile conditions. The document outlines the five main stages of micropropagation: preparation, initiation of cultures, multiplication, rooting, and transplantation. It provides details on each stage, focusing on choices of explants, factors influencing successful culture initiation, and methods of multiplication like regeneration from callus or direct shoot formation. Micropropagation offers advantages like high multiplication rates, disease elimination, and cryopreservation of plant materials.
Plant tissue culture involves growing plant cells, tissues or organs in sterile conditions on a nutrient medium. It allows for clonal propagation, production of secondary metabolites, induction of genetic variability, and regeneration of plants from somatic embryos or protoplasts. Key applications of plant tissue culture include micropropagation, production of pharmaceuticals, haploid production, somatic hybridization, transgenic plant production, germplasm conservation, breaking seed dormancy, and biomass energy production.
Commercial Exploitation of Micro-propagation in fruit crops & its TechniquesPawan Nagar
Micropropagation is a tissue culture technique where plantlets are regenerated from small plant parts like shoot tips, nodes, and meristems. It allows for the rapid multiplication of plant materials in a relatively short period of time compared to traditional propagation methods. The process involves sterilizing and culturing explants on nutrient media, multiplying shoots through subculture, rooting the shoots, and acclimatizing the plantlets. Micropropagation has various advantages like producing disease-free plants, conserving germplasm, and facilitating the export of plants. It has been commercialized for many horticultural crops in India like banana, citrus, grapes, guava, papaya, and strawberry through research institutes.
This document discusses several applications of plant tissue culture, including clonal propagation to produce genetically identical copies of plants, synthetic seed production, somatic hybridization to create hybrids between sexually incompatible plants, and cryopreservation for long-term germplasm storage. Clonal propagation is used in horticulture and forestry to multiply desirable traits from a single donor plant. Synthetic seeds encapsulate somatic embryos to allow for disease-free propagation of asexually reproducing plants. Somatic hybridization uses isolated protoplasts to create hybrids and introduce traits from wild plants to crops. Cryopreservation stores plant tissues and organs at ultra-low temperatures in liquid nitrogen for long-term conservation of plant germplasm.
Micropropagation is a method of rapidly multiplying plant materials through tissue culture techniques. It involves four main stages: selection and sterilization of explant material, multiplication of propagules in culture, regeneration of plantlets, and acclimatization of plantlets to soil conditions. The method allows for large-scale production of disease-free plants in a controlled environment and offers advantages such as year-round production and transport of materials across borders. However, it also has limitations such as high costs, need for specialized facilities and skilled personnel, and lack of confirmed genetic stability in some methods.
Plant tissue culture provides several benefits for studying plant growth and development. It allows scientists to isolate plant parts and culture them in vitro, simplifying the study of controlling influences. Some key applications of plant tissue culture include clonal propagation of disease-free plants, studying plant cells' ability to regenerate whole plants from cultured cells, producing genetic variability through somaclonal variation, regenerating plants from pollen to create haploids, and rescuing hybrid embryos. Plant tissue culture also enables fundamental biological research, production of high-value biochemicals, and generation of transgenic plants.
Plant tissue culture is used to produce valuable secondary metabolites. There are three main methods: cell suspension cultures, hairy root cultures, and immobilized cell cultures. Cell suspension cultures are the most common. They involve transferring plant cells or callus to liquid medium to grow as a suspension. Medium components, regulators, precursors, elicitors, and environmental factors can be manipulated to enhance metabolite production. Hairy root cultures use plant cells transformed by bacteria to produce root-like structures that also synthesize metabolites. Immobilized cell cultures entrap or affix cells to allow contact while protecting from shear stress. Each method aims to produce metabolites through optimized bioprocessing conditions at large scales.
This document discusses plant tissue culture techniques. It begins by explaining how plant tissue culture has become popular for horticultural and industrial applications as well as studying plant growth and development. It then describes the history and development of plant tissue culture, including its earliest uses and current commercial applications like micropropagation. The rest of the document details the various techniques involved in plant tissue culture, including quantitative and qualitative improvement approaches, micropropagation stages and methods, and troubleshooting browning issues.
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1. PLANT TISSUE CULTURE
AND ITS
APPLICATIONS
BIOLOGY
PROJECT
SUBMITTED BY
NAME: VAISHNOVI SEKAR
CLASS: XII C
ROLL NO:
2. INDEX
# AIM
# CERTIFICATE
# ACKNOWLEDGEMENT
# DECLARATION
# OBJECTIVE
# INTRODUCTION TO PLANT TISSUE CULTURE
# APPLICATION OF PLANT TISSUE CULTURE
# PLANT TISSSUE CULTURE FOR MASS PRODUCTION OF HIGH VALUE
SECONDARY METABOLITIES
# THEORY
# EXERCISE 1: PREPARATION OF STOCK SOLUTION
MACRONUTRIENTS STOCK SOLUTIONS
MICRONUTRIENTS STOCK SOLUTION
VITAMIN STOCK SOLUTION
IRON STOCK SOLUTION
# EXERCISE 2: PREPARATION OF MURASHIGE AND SKOOG(MS)
MEDIUM
# EXERCISE 3: INOCULATION OF EXPLANTS NODAL SEGMENT
# EXERCISE 4: PREPARATION OF MS MEDIUM SUPPLEMENTED WITH
PLANT GROWTH HORMONES FOR REGENERATION OF
EXPLANTS
# RESULT AND OBSERVATION
# BIBLIOGRAPHY
4. CERTIFICATE
This is to certify that this project is submitted by
VAISHNOVI SEKAR to the biology department,
CHINMAYA VIDYALAYA, NEW DELHI, was carried out by
her under the guidance and supervision of
Mrs Anubha Srivastav during academic session 20102011.
Date: 03 Nov. 10
Mrs Anubha Srivastav
(Biology teacher)
5. ACKNOWLEDGEMENT
I wish to express my deep gratitude and sincere
thanks to Mrs Anubha Srivastav for her
encouragement and for all the facilities that
she provided for this project work. I sincerely
appreciate this magnanimity by taking me into her
fold for which I shall remain indebted to her. I also
thank our lab assistant, Mrs. Nikhila Nair.
I take this opportunity to express my deep sense of
gratitude for her invaluable guidance, constant
encouragement, constructive comments,
sympathetic attitude and immense motivation in
all stages of this project work.
VAISHNOVI SEKAR
6. DECLARATION
I do hereby declare that this project work has
been originally carried under the guidance
and supervision of
Mrs Anubha Srivastav
Biology teacher
Chinmaya Vidyalaya
VAISHNOVI SEKAR
7. OBJECTIVE
The purpose of the experiment is – to grow new
plant segment with the help of invitro culturing of
the plant nodal segment. I got interested to this
project after learning about various technical
advancements in the field of growth and metabolic
engineering of plants.
Micro propagation of plants of medical importance
and multiplication of high yielding plants is done by
use of plant tissue culture and they are adapted to
local atmospheric conditions by biotechnological
approaches.
8. Introduction: plant tissue culture
Plant tissue culture or the aseptic culture of cells, tissues,
organs and their components under defined physical and
chemical conditions in vitro, has become am important
tool in both basic and applied research and in commercial
applications. It is an outcome of the genesis of an idea of
the German scientist, Haberlandt, at the beginning of the
20th century who believed in the concept of Toti potency.
It relies on maintaining plant cells in aseptic conditions on
a suitable nutrient medium. The culture can be sustained
as a mass of undifferentiated cells for an extended period
of time, or regeneration into whole plants. Designing a
strategy to culture cells from a plant for the first time can
still seem like a matter of trial and error, and luck.
However, the commercial production of valuable
horticulture crops by micro propagation shows that it
exists in the routine, as well as experimental world.
9. Applications: Plant tissue culture
Plant tissue culture now has direct commercial applications
as well as value in basic research into cell biology, genetics
and biochemistry. The techniques include culture of cells,
anthers, ovules and embryos on experimental to industrial
scales, protoplast isolation and fusion, cell selection,
meristem and bud culture.
Application include
Micro propagation using meristem and shoot culture to
produce large numbers of identical individuals
Large scale growth of plant cells in liquid culture as a
source of secondary products
Crossing distantly related species by protoplast fusion and
regeneration of the novel hybrid
Production of dihaploid plants through pollen or anther
culture to achieve homozygous lines more rapidly for
breeding programmes
As a tissue for transformation for regeneration of
transgenic plants
Removal of viruses by propagation from meristematic
tissues
Secondary metabolites production
Triploid through endosperm culture
Tissue culture offers an additional or alternative way of
propagation of selective genotypes. This technique is being
commercially exploited for mass propagation of
herbaceous, ornamental and fruit trees.
10. High value secondary metabolites
Plant tissue culture forms the backbone for plant
biotechnology. It encompasses micro propagation,
induction of soma clones, somatic hybridization, cry
preservation and regeneration of transgenic plants.
Interest in medicinal plants as a re-emerging health
aid has been fuelled by the rising costs of
prescription drugs in the maintenance of personal
health and well being, besides the reported side
effects and inability to cure chronic diseases, coupled
with bio-prospecting of new plant derived drugs. In
order to have standardised formulations, the
chemical constituents from plants and their parts are
required to be uniform both qualitatively and
quantitatively.
Furthermore, an ever increasing demand of uniform
medicinal plant-based medicine warrants their mass
cloning through plant tissue culture strategy. A good
number of medical plants have been reported to
regenerate in vitro from their various parts.
11. Most secondary metabolites are important medicinal
compounds. Plants cells culture provides an excellent
system for large scale production of these
compounds. There are a wide variety of secondary
metabolites such as taxol, artemisinin, morphine,
quinine, vinblastine, atropine, scopolamine and
digoxin that can be produced from plants. This
technique represents an optimal solution that has
negated problems such as the slow growth cycle of
the plant source. Plant cell culture allows for the
synthesis of the bioactive substances present in
plants, and is often the only available source,
unhampered by quantitative limitations, of active
ingredients which are poorly available or difficult to
manufacture by chemical synthesis.
Tissue culture is an alternative way for the
production of photochemical of therapeutic
importance. The technique like hairy root culture,
biotransformation, immobilization and elicitations
are used for the increased production of secondary
metabolites. By this approach, therefore, an
increased production of secondary metabolites can
be obtained.
12. Theory:
Plant tissue cultures can be initiated from almost any
part, the explants, of a plant. The physiological state
of the plant does have an influence on its response.
The mother plant must be healthy and free from
obvious sign of disease or decay. Younger tissue
contains a higher proportion of actively dividing cells
and is more responsive to culture conditions.
The exact conditions required to initiate and sustain
plant cells in culture, or to regenerate plantlets from
cultured cells, are different for each plant species.
Each variety of a species will often have a particular
set of culture requirements. Despite all the
knowledge that has been obtained about plant tissue
culture during the 20 th century, there conditions have
to be identified for each variety through
experimentation.
In tissue culture, propagation involves five important
steps:
Step I: involves the initiation of culture which should
be infection free. The culture may be of shoot, root,
nodal and inter-nodal segments etc. care should be
taken to ensure the survival of explants after
sterilization process.
Step II: is the rapid multiplication of culture to
produce plantlets regularly. In a number of species,
this stage is easily carried out by the axillary shoot
13. initiation or adventitious organ formation which is
preferred because somatic embryogenesis has been
utilized to produce faster seedling production. In
case of axillary multiplication, plants have the same
genetic pool as the maternal source. The rate of
multiplication is not high.
Step III: prepare the plant propagules for transfer
from in vitro or in vivo condition. It requires the
induction of roots and hardening of plantlets. Both
the processes start when they are maintained in
cultural conditions and receive in vitro treatments.
Step IV: transfer of tissue culture raised plantlets to
the field to adapt to natural environmental condition.
A number of methods have been applied which
includes the use of mist or fog, carbon-dioxide
enrichment, optimization of light and temperature,
etc. the transferred plantlets grow slowly at initial
stages and restore normal growth later.
Step V: the selection of mother plants is very
important for commercial success of tissue culture
propagation. It may involve the process where the
mother plants are grown under strictly controlled
conditions-25°C temperature with humidity of 75.
14. EXERCISE 1: STOCK SOLUTION
AIM: PREPARATION OF STOCK SOLUTION
PROCEDURE: PREPARE SEPARATELY THE STOCK SOLUTION OF
MACRONUTRIENTS, MICRONUTRIENTS, VITAMIN AND IRON BY
DISSOLVING ALL THE CONSTITUENTS INDIVIDUALLY IN CONICAL FLASK
OVER A MAGNETIC STIRRER.
THE COMPOSITION OF EACH STOCK IS GIVEN BELOW:-
Nutrients
Major nutrients
concentration
NH4NO3
CaCL2
MgSO4
KNO3
KH2PO4
Minor nutrients
16.5g in 100ml
4.4g in 100ml
3.7g in 100ml
19.0g in 100ml
1.7g in 100ml
H3BO4
MnSO4
ZnSO4
CuSO4
CoCl2
KI
Na2MoO4
0.124g in 100ml
0.312g in 100ml
0.172g in 100ml
0.025g in 100ml
0.25g
0.83g
in 100ml
0.25g
Iron solution
FeSo4
0.895g
Na2Edta
Vitamin
0.932g
Glycine
Nicotinic acid
Pyridoxine HCl
Thiamine HCl
0.1g
0.025g
0.025g
0.01g
in 250ml
in 100ml
Precautions:
Separately dissolve all salts to prevent any precipitation
Use lukewarm water for easy dissolution
16. Exercise 2: MS medium
Aim: Preparation of Murashige and Skoog medium
Procedure:
1. Required amount of major, minor, iron and vitamin stocks for preparation
of 1L of MS medium
Major stock
Minor stock
Iron stock
Vitamin
2.
100ml
50ml
10ml
2ml
Take 1L beaker
Dissolve 3% sucrose in 300ml double distil water
Add required amount of major, minor, iron and vitamin stocks
Make up the volume to 1L
Adjust pH to 5.70
(With the help of 1N HCL/1N NaOH)
Supplement medium with 0.63% agar
Boil the medium till agar gets dissolved
17. Pour the medium in culture vessels
(Test tubes/ flasks)
Plug the vessels, cover them with aluminium foil
Sterilize the medium by autoclaving at 121°C for 15 minutes
After autoclaving, the medium is kept inside the culture room.
Precautions:
the vessels should be sterilized in order to prevent the growth of
micro organism like fungus, etc.
18. Exercise 3: Inoculation
Aim: inoculation of explant, nodal segment
Procedure:
1. sterilization of explant
The nodal segments have to be treated before inoculation
a. under tap water for 15 mins
b. Tween 20 (2ml/100ml) for 20mins
c. Fungicide (Bavistin) (0.01g/L) for 45min
d. HgCl2(0.1%) for 10min
2. Preparation of laminar air flow chamber(LAFC)
Wipe the laminar air flow chamber with 70% alcohol.
Keep:
a. Spirit lamp (filled with spirit)
b. 1 flask containing absolute alcohol filled up to neck
c. Another flask containing 50ml (70% alcohol)
d. Match box
e. Sterile blades
f. Sterile water
g. Vessels containing medium
h. Sterile instrument pair
i. Sterile pertiplate
In the laminar air flow chamber
Switch on UV light in the LAFC for 15 minutes
19. Switch on Air flow after 15 minutes
Can start inoculation
3. Inoculation of explants
Wipe the hands with 70% alcohol
Fix the blade in the scalpel, flame the forceps and scalpel,
After flaming, keep them back in absolute alcohol
With the help of forceps, drop the explants into sterile water containing
flask
Flame the sterile Petri plate with 70% alcohol
Wait for 1-2 minutes so that the surface of pertiplate gets cooled
Take out the explants (nodal segment) from the sterile water
Cut from the top and bottom so that the sterilant at the two ends doesn’t
affect the result
Care should be taken to maintain polarity
Hence cut the lower end can be cut in slanting way so that it can be
demarcated
20. Open the vessel containing medium in front of the flame
Pick the nodal segments and insert the lower cut end into the medium
Close the vessel by putting back the plug in front of the flame
Keep the culture in the culture room
Take observation periodically
Culture room conditions
Temperature: 25±2°C
Humidity: 55±5%
Photoperiod (light/dark):16hr/8hr
21.
22. Exercise 4: MS medium
supplemented with plant growth
hormones
1.
Preparation of stock solution
Weight 50mg BAP and put it in sterile flask
Dissolve BAP in 1N HCL (2ml)
Make up the volume up to 50ml with the help of double distilled water
Store the stock in transparent glass bottle and label the content
(1mg/ml)
Keep it in refrigerator at 20°C
2.
PREPARATION OF MEDIA (500ml)
2.5mg/l
Take 2L beaker
Dissolve 15g sucrose in 500ml double distilled water
(3% sucrose is used)
Add required amount of major, minor, iron and vitamin stocks
23. Make up the volume to 500ml
Supplement medium with 0.63% agar
Pour the medium in culture vessels, label them A-F
Plug the vessels, cover them with Aluminium foil
Sterilize the medium by autoclaving at 121°C for 15 minutes
After autoclaving, the medium is kept inside the culture room
Inoculate the explants on above medium after 48 hours of media
autoclaving
Study regeneration process periodically
26. Conclusion:
Micro-propagation is the development of new plants in
an artificial medium under aseptic conditions. This
method is employed for rapid multiplication of elite
plant species.
Discussion:
The most frequently used micro propagation method for
commercial production utilizes enhanced axillary shoot
proliferation from cultured meristem.
Micro propagation process supports in achieving mass
production of healthy plants in low or minimum space
requirement.
On the other hand, micro propagation process comprises
of high labour costs, danger of variation and loss by
contamination.
Multiplication rates are tend to be slow at first, but
later, if the cultural conditions are satisfactory (temp:
25 ± 2 °; humidity 55± 5%; photoperiod – light/dark:
16 hrs/8 hrs), a rapid multiplication can be achieved.
Generally, the plant taken for micro propagation are the
ones which are high in demand in terms of number, are
in demand round the year and have to be multiplied
irrespective of season, are amongst the endangered
ones, have to be cloned, or have to be preserved for
their special qualities.