1. What are stem cells? What are the types and applications of stem cells?

Stem cells Stem cells are progential cells which have the ability to change into many different types of cells by differentiation and specialization. Types of stem cells: 1. Embryonic stem cells (ESC): 4 to 5 days after the formation of Zygote, it develops into a mass of cells surround by single layer of cell known as Blastocyst. The inner mass cells of the blastocyst have the ability to differentiate and develop into any of the 250 different types of cells in human body. Such progential embryonic cells are known as embroyonic stem cells. The ability of ESC to differentiate into all kinds of cells is known as Tatipotency. 2. Fictal stem cells: In the foetus certain cells have found to have stem cell property and are known as fictal stem cells (FSC). Their ability to differentiate to many types of cells but not all is called Pluri potency. 3. Adult stem cells Certain cells in adult body differentiate into few kinds of cells as adult stem cells. Their ability to differentiate into very few kind of cells is known as Multi potency. Eg: Bone marrow cells, limbus cells of eye, oral mucous cells. 4. New types of stem cells: a) Cord blood stem cells: The cells present in umblical cord that can be stored for future cells. b) Amniotic fluid derived stem cells ( AFDSC): Certain foetal cells have been found to have stem cells properties.

Applications: The major application of stem cells is regenerating the damaged parts in various diseases. The branch of medicines that uses stem cells for this procedure is called Re-generative medicine. Using embryonic stem cells in regenerating the damaged part is known as Therapeutic cloning. The principle used in therapeautic cloning is somatic call nuclear transfer. Using this many types of diseases can be cures which include Type-I diabetes, types of cancer, Alzemiers, Parkinson disease, spinal injuries, arthritis etc. 2. What do you know about stem cells? The ancestral cells, which can divide and give to progeny that undergo differentiation are called stem cells. Zygote is a totipotent cell. It can give rise to the whole organism. Pleuripotent cells arise from totipotent cells. They give rise to most, but not all types of cells. Embryonic inner cell mass( embryonic stem cells) of blastocyst contains pleuripotent cells. Multipotent cells are differentiated from pleuripotent cells. Multipotent cells can give rise to only a limited number of cell types. Each unipotent cell gives rise to a single modified cell type. 1. Embryonic stem cells: Embryonic stem cell lines (ES cell lines) are cultures of cells derived from the epiblast tissue of the inner cell mass of a blastocyst. ES cells are pluripotent cells and give rise three primary germ layers-ecoderm, endoderm and mesoderm. They can develop into more than 200 cell types of the adult body. When given no stimuli for differentiation, ES Cells will continue to divide invitro and each daughter cell will remain pluripotent. 2. Adult stem cells: Adult stem cells are undifferentiated cells found throughout the body that divide to replenish dying cells and regenerate damaged tissues, also known as somatic stem cells. Haemopoietic stem cells (HSCs) are pluripotent and multipotent cells. These are also the self renewing cells.

These HSCs produce two secondary stem cells( common progenitors) namely myeloid stem cell and lymphoid stem cell. These are nonrenewing cells. i) Cells formed from the Myeloid stem cell: In the erythroid lineage, the erythrocyte is formed through different stages like proerythroblast, erythroblast and reticulocyte. Megakaryocyte forms into platelets. Basophil and eosinophil committed progenitors from the basophil and eosinophil respectively. Monocyte is formed from monoblast and neutrophil is formed from mycloblast committed progenitor. Megakaryocyte is formed from megakaryoblast. ii) Cells formed from the Lymphoid stem cell: Lymphoid common progenitor gives rise T-cell committed progenitor and B-cell committed progenitor. It also gives rise to natural killer cell and dendritic cell directly. Some stem cells remain in the epithelium of skin and gastrointestinal tract to replenish the last cells of apical layers. Skeletal muscle tissue contains the stem cells called satellite cells to replace the damaged striated muscle cells. Smooth calls contain the pericytes to replace the damaged smooth muscle cells. 3. Application of stem cell research: Transplantation of HSCs to the patients is an important application. Human embryonic stem cell (ES cells) are used to replace deceased or damaged cells in the brain and in treating cell based diseases like Parkinson disease. The best known stem cell therapy till date is the bone marrow transplant, which is used to treat leukemia and other types of Cancer as well as various blood disorders. The multipotent stem cell rich blood found in the umbilical card is useful n treating the above disorders.

3. What are transgenic transgensis ?

animals?

What

are

the

methods

of

Animals that have their DNA manipulated to possess and express on extra (foreign) gene are known as transgenic animals. Transgenic rats, rabbits, pigs, cows and fish have been produced although over 95% of all existing transgenic animals are mice. In creating a transgenic animal, the transgene should be introduced just after fertilization into the male pro-nucleus known as transgensis. There are 3 methods of transgensis. 1. Micro Injection 2. Nuclear Transplant 3. Trans-embryo technology. 1. Micro-Injection: It involves introduction of trans-gene into the male pro-nucleus of the zygote just after fertilization, by a microscopic needle. The probability of gensis is 10%.

2. Nuclear transplant: This method involves introducing a transgene into a separate nucleus and then transplanting it into a e-nucleated egg. The probability of transgensis is 2.5%.

3. Trans-embryo technology: This method involves the use of lenti-virus to insert transgene into the male pro-nucleus. Viruses that integrate their genetic material with host cell genome are known as lenti viruses. 100 % gensis is possible by this method. The most widely used technique to produce transgenic organisms is micro-injection since it is very easy to carry out.

4. What are transgenic animals? What are their uses/applications? Animals that have their DNA manipulated to possess and express on extra (foreign) gene are known as transgenic animals. Transgenic rats, rabbits, pigs, cows and fish have been produced although over 95% of all existing transgenic animals are mice. Some of the benefits from such modifications are: 1. Normal physiology and development: Transgenic animals can be specifically designed to allow the study of how genes are regulated and how they affect the normal functions of the body and its development. E.g. study of complex factors involved in growth such as insulin like factor. By introducing genes from other species that alter the formation of this factor and studying the biological effects that result information is obtained about the biological role of the factor in the body. 2. Study of disease: Many transgenic animals are designed to increase our understanding of hoe gees contribute to the development of disease. They are specifically made to serve as models for human diseases so that investigation of new treatments for diseases is made possible. Today transgenic models exist for many human diseases such as cancer, cystic fibrosis, rheumatoid arthritis and Alzheimers. 3. Biological products: Medicines required to treat certain human diseases can contain biological products, but such products are often expensive to make. Transgenic animals that produce useful biological products can be created by the introduction of the portion of DNA( or genes) which codes for a particular product such as human protein( -1- antitrypsin) used to treat emphysema. Similarly attempts are being made for treatment of phenylketonuria (PKU) and cystic fibrosis. In 1997, the first transgenic cow, Rosie, produced human protein-enriched milk(2.4 gma/litre). The milk contained the human alpha- lactalbumin and was nutritionally a more balanced product for human babies than natural cow milk.

4. Vaccine Safety: Transgenic mice are being developed for use in testing the safety of vaccines before they are used on humans. Transgenic mice are used to test the safety of the polio vaccine. If successful and fund to be reliable, they could replace the use of monkeys to test the safety of batches of the vaccine. 5. Chemical safety Testing: Transgenic animals are made that carry genes which make them more sensitive to toxic substances than non transgenic animals. They are then exposed to the toxic substances and the effects studied. Toxicity testing in such animals will allow us to obtain results in less time.

5. What are the ethical implications on transgenic animals? 1. Creation of new life forms: i) The transfer of genes between two species may in course of time affect the genome of the species, leading to damage in the original genome. Transgenic animals may interact with the environment in a way which is not likely for the sustainability of safe environment and produce unexpected outcomes. Changing genomes of animals may lead to reduction in breeding ability and life span.

ii)

iii)

2. Increased use of animals: i) While constructing transgenic animals, the gen may get integrated at any site of the genome of the host cell. So slightly difference phenotypes are expressed. Besides this, many transgenic individuals are required to analyse the correct phenotype of the transgenic animal. In transgenic disease models, one model has not been used for other diseases or even other workers, as it has the selective target gene

ii)

alone. So the model animals have to be killed. This is an animal wastage. 3. Opportunities for Reduction of Animal Use: i) Transgenic animals are better models than conventional animal breeds for study of genes, bio processes and diseases. A few animals are enough to collect complete statistical data. Transgenic animals in some cases seem to be substitute for human models.

ii)

4. Creation of unexpected phenotypes: The genetic and physiological characters in most cases of transgenic animals are found different from that which were expected. This is mainly due to unexpected interaction of the desires gene with other genes in the genome (this is one of the most common problem in mice). 5. Pathological Effect: 1. Uncontrolled expression of inserted gene is noticed in several transgenic mice due to over expression of the gene. It may increase the morbidity and mortality. Transgenic animals with inserted growth harmone gene are examples of this problem. 2. For constructing transgenic models, normal functional gene of animals is replaced with non functional gene. So the normal genes become ineffective.

6. Write about Transgenic plants.

The novel plants with desirable characters created through genetic engineering methods are called transgenic plants. The production of transgenic plants has become a routine practice in many laboratories. Initially, the production of transgenic plants was restricted to dicotyledons but it has now been extended to several monocotyledons like wheat, rice, maize, Oats etc.

Beneficial aspects of transgenic plants: 1. Transgenic plants are important in increasing the efficiency of crop plants. For instance, transgenic plants resistant to herbicides, insects and viruses have already been produced. 2. Transgenic plants have also been produced, which are suitable for food processing. Bruise resistant and delayed ripening tomato plants are produced by transgenics. 3. Male sterile plants of Brassica napus are produced. This will eliminate the problem of manual emasculation and controlled pollination to reduce the cost of hybrid seed production. 4. Transgenic plants can be used as bioreactors for obtaining commercially useful products, specialized medicines, chemicals and antibodies for large scale production. This area is described as molecular farming. 5. Transgenic plants have been produced for identification of regulatory sequences for many years.

Inspite of impressive progress in plant transgenics, the actual commercialization of the transgenic crop plants has been quite slow. The main reason is due to apprehension from the public towards biosafety issues of genetically engineered crops.

7. What are Transgenic plants? Plants containing introduced DNA, are known as transgenic plants or genetically engineered plants. They have acquired a new trait from the introduced DNA ad inherit the trait of many generations. In some transgenic plants, the introduced DNA blocks the normal functioning of certain original genes of the plants. Scientists have developed transgenic plants such as 1. Herbicide-resistant plants: These plants tolerate the herbicide and are safe in the field, when they are applied. e.g Glyphosate resistant petunia, tobacco, tomato and corn etc.,

2. Insect resistant plants: These plants have been developed by adopting gene transfer methods. The gene for Bt.-toxin was isolated from Bacillus thuringiensis, linked with 35 S promoter of Ca M V and introduced into plant using Ti plasmid as vector. The Transgenic plants developed from these cells are resistant to many insect pests. e.g: Bt-Cotton, Bt-Soya etc. 3. Virus-resistant plants By genetic engineering, plants are made to have parts of viral protein coat or virus nucleoproteins. The viral coat protein or nucleo protein interferes with infection by the related viruses. Some transgenic virus resistant plants are mentioned below: Crop Tobacco Tomato Alfalfa Cucumber Rice Papaya resistant to Tobacco streak virus (TSV) Tomato spotted wilt virus (TSWV) Alfalfa mosaic virus (AMV) Cucumber mosaic Virus (CuMV) Rice stripe Virus (RSV) Papaya ring spot virus

4. Transgenic plants with improved storage proteins Like French bean and transgenic potato, these are nutritionally valuable as they are rich in starch and amino acids. These transgenic plants can correct protein deficiency malnutrition in man. 5. Transgenic plants with improved oils and fats: The transgenic rapeseed is rich in sterates and oleic acids. The stability of its oil during frying is high and here is no problem of cholesterol. The transgenic soyabean yields cocoa oil that can be used in chocolate making.

6. Male sterile plants : These plants help breeders for hybridization, especially to avoid emasculation. The transgenic plants regenerated from the calli are male sterile. 7. Transgenic plants with attractive flowers: Attempts have been taken to change the flowers colours by introducing certain genes involving in flavoxoid metabolism or ontisense RNA producing genes. 8. Stress Tolerant Plants: Genetic engineers have developed new strains of plants with stress tolerance due to extreme cold, hot and drought. E.g:- In transgenic tobacco, due to the introduction of glycerol-iphosphate acyl transferase, mannitol dehydrogenase and sorbital dehydrogenase ans superoxide dismutase genes, it is resistant to chilling as low as 100, drought and to high light intensity. 9. Engineering for preservation of fruits: Transgenic plants with bruise resistance and delayed fruit ripening have been developed. Eg: Transgenic tomatoes produce aminocyclopropane carboxylic acid (ACC) for delayed fruit ripening. 10. High rate of photosynthesis : Mutant RUBISCO with low oxygenase activity was isolated from different plants and introduced into tobacco ealli which showed higher rate of photosynthesis. Research is going on to increase photosysthesic efficiency of various crop plants. 11. Transgenic plants as Bio reactors: Transgenic plants are used for the large scale production of some valuable products such as interferons, vaccines, biodegradable plastics etc.,

8. What is Biomedical Technology? Biomedical Technology involves the application of engineering and technology principles to the biological systems. Biomedical engineering combined with Bio-technology is often called Biomedical Technology or Bioengineering. Diagnostic imaging like X-ray radiography, computerized axial tomography and magnetic resonance imaging are some of the diagnostic tools. The x-ray technology is most commonly employed for diagnostic imaging. x-rays are useful in the detection of pathology of the skeletal system, detecting some disease processes in soft tissue etc. other imaging alternative for soft tissues are computed axial tomography, magnetic resonance imaging(MRI) etc. X-rays are listed as a carcinogen. An MRI scan is a radiology technique that uses magnetism, radio waves and a computer to produce images of body structures. The electrocardiography is a non invasive test that is used to detect heart conditions by measuring the electrical activity of the heart. An EEG is a test that can help diagnose epilepsy with seizures (emotional fits) and dementia. During an EEG the electric signals of the brain are recorded.

An organ transplant is the moving of a whole or partial organ from one body to another for the purpose of replacing the recipients damaged organ with a working one from the donor site. The transplantations may be life saving or life enhancing. The grafts are autograft, isograft, allograft and xenograft.

ELISA is an enzyme linked immune sorbent assay test. It requires an antibody and a substrate (an enzyme). The most basic type of ELISA consists of an affinity for the substance of interest, for example, human chorionic gonadotrophic (HCG), the commonly measures protein which indicates pregnancy.

9. Give an account on the scope and application of bio technology? Biotechnology is the science of utilizing the properties and uses of microorganisms or to exploit cells and the cell constituents at industrial level for generating useful products essential to life and human welfare. Some of the areas which have been utilizing the biotechnological methods for the production of specific compounds as: 1. Antibiotics are produced by microorganisms like actinomycetes, bacteria and fungi. Tremendous improvements were made in the process of development and productivity with the help of biotechnological advances. For instance, today it became possible to produce penicillin at a cost less than 1% of its cost production in 1945. 2. Biotechnology helped us to create and commercially produce pharmaceutical drugs like anticancer agents, interferons, human insulin and growth harmones. 3. Sugar substituting sweetners are being biotechnological process and marketed out. produced based upon

4. Biotechnology has also helped to increase the commercial production of enzymes. High fructose corn syrups (HFCS), the artificial sweetners are being produced on a mass scale using immobilized glucose isomerase. 5. A number of food supplementing agents like Monosodium glutamate (flavouring agent), aspartate, nucleotides and lysine are produced in large amounts to supplement foods and animal feeds. 6. The rennet used for the commercial production of cheese is produced by using genetically engineered microorganisms. 7. Biotechnology has opened up new avenues for the production of proteins from non-conventional sources like microorganisms. These microbial proteins are often referred to as single cell proteins(SCP). 8. In the area of plant protection, Bacillus thuringiensis and baculo viruses like NPV(Nuclear Polyhedrosis Virus) are commercially produces and used as biopesticides. Similarly many improvements were made in the production of biofertilizers.

10. Write an account on Bio-fertilizers. Bio fertilizers denotes the nutrient input for plant growth which is of biological origin. Biofertilizers comprise of a variety of living organisms. The nitrogen produced by nitrification, ammonification, nitrogen fixing bacteria, solubilizers of rock phosphate and other microorganisms which help in increasing the fertility of soil can be called as bio fertilizers. Some of the bio-fertilizers are given below: a) Rhizobium inoculants: Rhizobium inoculants are being increasingly used to enhance the productivity of legume crops. Rhizobium inoculants are applied to the legume seeds with gum or carboxy methyl cellulose and the seeds are then sown in the field. The plants which develop from such seeds possess larger number of root nodules with better growth and yield. b) Azospirillum: In Maize, Sorghum, wheat, barely and finger millet, Azospirillum lives as an associated symbiont. Besides the ability to fix atmospheric nitrogen, Azospirillum also secrets certain growth promoting substances. The technique of seed-dressing with bacteria ( e.g Azobacter, Azospirillum, Rhizobium etc.) is called Bacterization. They are multiplied in artificial media to harvest on a large scale for supply to the farmers. c) Cyanobacteria: (Blue green algae) such as Anabaena and Nostoc either independently or in symbiotic association fix atmospheric nitrogen. Farmers can develop cyanobacterial noculants in their fields by following simple procedures. These inoculants develop as dense mats which are to be cut into small fragments or blocks and can be introduced into rice fields. d) Azolla: Azolla is a water fern. In Azolla leaves, the blue green algae Anabaena azolle lives as an endosymbiont and fixes nitrogen. Azolla adds 30-40 kg of nitrogen per hectare. By using Azolla as bio-fertilizer in rice fields, yield can be increased. e) Mycorrhizae: The association between fungal members and roots of vascular plants is called mycorrhizae. Such roots associated with a fungus are called mycorrhizal roots. Arbuscular mycorrhizae (AM) fungi increase the phosphate absorption by the roots. Researchers revealed

an increase in the growth and yield of Potato, wheat, maize, Soybean and red gram by the employment of AM fungi e.g: Glomus. The use of AM inoculum improves plant growth, yield and increases resistance against environmental stresses and pests. Importance: The use of bio-fertilizers offers several advantages. They include: 1. By the application of bio-fertilizers, the use of harmful chemical fertilizers can be curtailed substantially. 2. There is no environmental pollution problem. They are eco- friendly in nature. 3. Bio-fertilizers are economical. By following some simple procedures, even the farmers can develop bio-fertilizers on their own. 4. Bio-fertilizers production requires no energy input in the form of fossil fuels. 5. Crop produce will not contain any toxins. 6. Some bio-fertilizers are useful not only to increase the growth and yield but also increase the resistance against environmental stresses and pests. 7. Some bio-fertilizers are useful not only to increase the growth and yield but also increase the resistance against environmental stresses and pests. Limitations: 1. Bio fertilizers are not developed to a stage where they can be reproduced en-moss and supplied to all farmers. 2. Even though they are good substitute for chemical fertilizers, they are not a complete alternative for chemical fertilizers. 3. The bio-fertilizers that are in use currently are not applicable to all types of crops. 4. They give good results only in definite soil conditions.

11.

Write an account on Bio-pesticides.

Bio-pesticides: The microorganisms used to control insects, pathogens or weeds that affect the crop plants are called Bio-pesticides. Biopesticides are classified into bio herbicides and bio insecticides. a) Bio herbicides: These are organisms or their extract which destroy weeds without harming useful plants. Sunflower, Soyabean, alfalfa, sweet clover etc., act as smooth crop and do not allow weeds to grow. b) Bio insecticides: living organism which are used to kill or repel insects are called bio insecticides. There are three types of bio-pesticides. 1. Microbial pesticides 2. Botanical Pesticides 3. Bio-chemical pesticides. 1. Microbial Pesticides: Microorganisms used for bi control are bacteria, viruses, fungi and protozoa. a) Bacteria as bio-pesticide: A number of bacteria infect but only a few species in the genus Bacillus received attention for their use as bio pesticides. Among the species of Bacillus the most important one for its use as bio-pesticides is Bacillus thuringiensis (Bt). Other species are B.Popilliae, B.Sphaericus and B.lentimorbus. b) Viral Pesticides: Amongst viruses only baculovirus group are considered are considered potential pesticide. Most popular are the nuclear polyhedrosis viruses (NPHV). These viruses are often highly virulent and quickly disseminate the insect population such as caterpillars and moths. c) Fungal Pesticides: Many fungi attack insets and cause disease in them. They are called entomopathogenic fungi, which are now used to control the pests and are known as fungal pesticides. Eg: Beaveria bassiana.

2. Botanical pesticides: Many plants extracts have insecticidal property. So they are directly extracted from proper plant species and sprayed over crops to control insect pests. The plant extracts that control pests are named botanicals or botanical insecticides. Eg: a) Azadirachlin is an alkaloid present in Azadirachta indica (Neem tree). Neen extracts control about insect pests, many nematodes and pathogens of crops. b) Black leaf-40 is a spray- mixture formulated by mixing tobacco extract and copper sulphate. It is more toxic to insects and even human beings too. c) Pudina extract from leaves of Mentha Piperata and Mentha Arvensis. It is used to control the germination of fungal spores on stored grains. 3. Bio-chemical Pesticides: Bio-chemicals control pests by harmless mechanism. The most widely used bio-chemical pesticides are pheromones ( insect sex pheromones). They act as sex attractants and help in trapping the male and female insects that damage crops. Advantages: 1. Bio pesticides are biodegradable, safe and less stable. 2. They are very cheap. 3. They do not give residual effects in plant products. 4. They are usually more specific in action. 5. They do not have any effect on soil micro-life. 6. They do not persist for a long time without their proper hosts. 7. Bio-pesticides if used as a part of Integrated Pest Management (IPH) can reduce the dependence on chemical pesticides.

12. Write an account on biofuels The inflammable substances produced by the action of microbes, that can be set on fire are called Biofuels. They include ethanol, methane and hydrogen. They are being produced from cheap raw materials or wastes. They do not release high proportion of CO2. About 15% of the total energy consumption of the world is supplied by biofuels. Biofuels are of two types: Bio ethanol and Bio diesel. 1. Bio ethanol: Ethanol (Ethyl alcohol) is a colourless, inflammable liquid. It is spirituous in odour. It is also known as grain alcohol, spirit and gasohol. It is a good solvent for fats, resins, dyes, etc. It can be used as a fuel for lamps and stoves and in running internal combustion engines. In biotechnology based industries, ethanol is produced from wastes by the action of microbes. Any waste rich in carbohydrate is used as the substrate to produce ethanol. 1st method 2 CH3COCOOH--------------- 2CH3CHO + 2CO2 CH3CHO + NADH2----------------------- CH3CH2OH + NAD 2nd Method: C12 H22 O11 + H2O------------------------- 2C6H12O6 C6H12O6 -------------------------2C2H5OH + 2 CO2 2. Bio diesel: Biodiesel is clean burning oil manufactured from fat or vegetable oils transesterification process. It is not of petroleum origin, but it can be blended with petroleum diesel for being used as a fuel. Biodiesel can be used in ordinary diesel engines with little or no modification. It is biodegradable, non toxic and free from sulphur and aromatics. Methyl esters of fatty acids are prepared by treating the fatty acids with an ethereal solution of diazomethane or Boron trifloride(BF3)

RCO2H + CH2N------------------RCO2CH3 + N2 The methylated fatty acid is then converted into methyl ester of fatty acid (biodiesel) and glycerol by transesterification or alcoholysis. R.CO2. CH3 + C2H5OH ========= R.CO2.CH3 + Gl.

Advantages : 1. Dependence on convention fuels like petrol/ diesel decreases. 2. Self- sufficiency in energy sector can be achieved. 3. Plantation of Jatropha and Pongamia increase green cover. 4. Increase in rural employment. 5. Marginal/waste land can be brought under cultivation.

13.

Write an account of Biodiesel

Biodiesel is a clean burning oil manufactured from fat or vegetable oils by transesterification process. It is not of petroleum origin, but is can be blended with petroleum diesel for being used as a fuel. Biodiesel can be used in ordinary diesel engines with little or no modifications. It is biodegradable, non toxic and free from sulphur and aromatics. Manufacture: Methyl esters of fatty acids are prepared by treating the fatty acids with an ethereal solution of diazomethane or Boron triflouride(BF3). R.CO2 H + CH2 N----------------- RCO2 CH3 + N2 The methylated fatty acid is then converted into methyl ester of fatty acid (biodiesel) and glycerol by transesterification or alcoholysis. The methylated fatty acid is mixed with excess of alcohol and small amount of sodium alkoxide as a catalyst in a large air-tight container. The alcohol splits the glycerol from the methyl fatty acid. The glycerol is separated with the alcohol fraction and the remaining methyl fatty acid is stored and used as the biodiesel.

R.CO2 .CH3 + C2 H5 OH ============= R.CO2 CH3 + Gl

The important advantages of biodiesel are: 1. Bio degradable Fuel: Bio diesel is quickly degraded into natural organic residues by microbes. So there is no chance for oil pollution in soil and marine environments. 2. Non- toxic fuel: Bio diesel is more toxic than table salt. If it enters the gut, it lubricates the digestive tract. It doesnt make cracks or dryness on the skin of users and mechanics. 3. Reduction in Green House Gas: Bio diesel when set on fire, releases 78% less CO2 than that of petroleum diesel. Besides this, the plants use excess of CO2. A gradual recycling of CO2 is done. 4. Reduction of Air Pollution: The emission coming from biofuels B20 contains 12.6% CO instead 43.2% in diesel. Its mutagenic compounds are only 20% but they are 80-90% in the emission of diesel. So, it is believed that biodiesel can reduce the cancer risks by 94%. The sulphur dioxide emission is very low. Biodiesel fraction in the B20 reduces the odour of petroleum diesel. 5. Storage: Biodiesel can be stored up to 6 months without any treatment. As it is not auto ignitable, it can be retained safely. Disadvantages: 1. Bio diesel is not cheaper than petroleum diesel. 2. During cold weather, biodiesel gets thickened. So, it must be heated gently before use.

14.

How is Bio processes?

technology

used

in

Environmental

Clean

up

Bioremediation is the cleaning up of toxic contaminants in the environment using the activity of natural microbial populations in the contaminants or other wastes. Microbes act as catalysts to reduce the level of hazardness of toxic chemicals. Bioremediation is done the following ways. 1. Composting: The process of degrading explosive wastes by adding them to a pile of decaying organic matter/compost is known as composting. Hexa hydro trinitro tri- azine (RDX) and octa hydro tetrazocine (HMX) can be degraded by anaerabic bacteria and trinitro toluene (TNT) by both anaerobic and aerobic bacteria. 2. Land farming : The piling and maintenance of soil sludge on a flat land to degrade oil wastes by microbes existing in the sludge, is known as land farming. It is done on a stretch of flat land with clay soil in order to prevent seepage of contaminated water. Drawbacks: a) The process is very slow and incomplete. b) The heavy metal constitutes of the sludge gradually accumulate in the land farm soil. c) After land forming, the lands cannot be used for growing crops or grzing live stock. 3. Above ground bioreactors: These are based on the same technology as fermenters. They are used for the treatment of either excavated soil or ground water containing high levels of contaminants. Microbial growth in the bioreactors leads to a rapid rate of biodegradation. 4. In situ bioremediation:

The stimulation of natural bio transformations by adding nutrients to the environment is called enhanced in situ bioremediation. It is cheaper than incineration and workers are not exposed to the risks associated with excavation and removal of contaminated soils. Microorganisms participate in oil spill cleanups by oxidizing the oil to carbondioxide. Volatile hydrocarbon fractions evaporate quickly, leaving behind longer chain aliphatic and aromatic components. For most largescale oil spills, a combination of both human and microbial efforts is needed to affect a satisfactory cleanup in a reasonable amount of time. 15. Write an account of Biodegradation.

The breakdown of toxic substances by treating with efficient strains of microorganisms is called Biodegradation. Pollutants such as mercury compounds, arsenic compounds and biocides are degraded in this way. Xenobiotics are man-made chemicals that are present in the environment at unnaturally high concentrations. Derivatives of heavy metals and biocides are examples of Xenobiotics. Some specific strains of bacteria can tolerate heavy metals or biocides and degrade them to lose their toxicity. These bacteria are called biodegrading agents. E.g. Staphylococcus aureus 1258 degrades mercury compounds. The bacterial strain corynebacterium flaccum faciens DGIOI has a plasmid PDGIOI that provides resistance against arsenic compounds. It takes in arsenic compounds along with phosphates and excretes it into the surroundings in a harmless form. Some bacterial strains contain plasmids that confer resistance against the biocides. E.g. Pseudomonas putida BS893 has the plasmid PBS 241 which confers resistance against Biphenyl compounds. Treatment of Toxic Pollutants: A properly modified inoculum is diluted and sprayed over the contaminated soil. Then inorganic fertilizers such as nitrates and phosphates are spread over the soil. The contaminated soil is made wet by spraying enough water. The bacteria grow well and degrade the pollutant. Hence the concentration of the pollutant comes down to a great extent.

In usual practice, a combination of two or more strains is used to treat contaminants in the soil. The combined effect is better than the effect given by any of the pure strains. Advantages: 1. Toxic substances that cannot be disposed biologically are detoxified in this way. 2. Biodegradation helps to detoxify the toxic effects of biocides and heavy metals in the soil and water. 3. It is cheap to work out. 16. Write critical account on biodegradation of industrial effluents containing heavy metals.

The effluents coming from tanneries, textiles, metal polishing industries, fertilizer factories etc. contain heavy metals such as As, Cd, Cr, Pb, Ni, Hg & Zn. The process of capturing of metal contaminants from waste waters using microorganisms is called Biosorption. Some microorganisms attract ions of heavy metals in liquids and capture them. They are called Biosorbents. Biosorbents capture heavy metals through active and passive processes. In the active process, microbes capture and uptake metal ions from waste water by using cellular energy. e.g. Hyphomicrobium concentrates Mn; Gallionella Sps concentrates Fe and Cu; Pseudomonas aeruginosa concentrates Ur, Cr, Mo, Ca and Pb. In the passive process, no cellular energy is involved. Heavy metals have a tendency to bind with proteins, lignin, chitin and cellulose derivatives on the cell wall. They move towards the cell components and stick with them. Saccharomuces cerevisiae and Rhizopus arrhizus accumulate Uranium on their cell wall by passive process. Method for removing Heavy Metals: Cells of a suitable bacterium that has the capacity to capture the particular heavy metal are immobilized on filters or in pellets and kept inside a large bioreactor.

Waste water to be treated is pumped into the bioreactor to carry out biosorption. The microbial cells capture the heavy metal ions from the water and reduce their concentration in the water. After a period of specific time, the waste water is left out of the reactor to discharge it into a river. As the waste water has least amount of heavy metals, it has no polluting effect on the river water. Advantages: 1. By using biosorption, 90% of Uranium in nuclear waste streams is reduced. 2. Heavy metal ions in industrial effluents are reduced to a considerable extent before discharging them into rivers. 3. It can be adopted in industries for continuous operation to reduce pollution hazards. 4. This method can be adopted for liquids containing mild concentration of metal ions. 17. How microbial biotechnology helps in controlling heavy metal pollutants?

Genetically modified microbes offer a cheap way to clean up heavy metal pollution. Although the toxic metals remain in the soil, once bound to the modified bacteria, it is less likely to be taken up by plants and animals. The cell surfaces of many microbes have receptors that bind to heavy metals, but this metal sequestering is too feeble to significantly reduce metal pollution in soil. Victor de Lorenzo and colleagues of the centro Nacional de Biotechnologia in Madrid and Spain have engineered a mouse metallathionein on the cell surface of Ralstonia eutropha, a bacterium natural resistant to heavy metal pollution. In tests using cadmium contaminated soil and tobacco plants, the modified bacteria bound 3 times as much cadmium as unaltered microbes, reducing the cadmiums toxic effect on plants. Over 4 times as many plants grew in treated as against untreated soil. Heavy metal-resistant bacteria have been studied for their potential in leaching metal rom ores and cleaning up contaminated industrial sites. Some strains of bacteria can accumulate metals in their cell walls, some times as much as a quarter of their dry weight. Bacteria belonging to the

genus pseudomonas, among others, have been isolated that can tackle mercury, nickel, cobalt and other metal contaminants in waste. 18. What are the biotechnological applications in agriculture?

Three options for increasing food production are: 1. Agro-chemical based agriculture; 2. Organic agriculture; and 3. Genetically engineered crop based agriculture. The green revolution succeeded in tripling the food supply but yet it was not enough to feed the growing human population. Increased yields have partly been due to the use of improved crop varieties, but mainly due to the use of better management practices and use of agrochemicals (fertilizers and pesticides). However, for farmers in the developing world, agrochemicals are often too expensive and further increases in yield with existing varieties are not possible using conventional breeding. Plants, bacteria, fungi and animals whose genes have been altered by manipulation are called Genetically Modified organisms (GMO). GM plants have been useful in many ways. Genetic modification has: Made crops more tolerant to abiotic stresses( cold, drought, salt, heat) Reduced reliance on chemical pesticides(pest-resistant crops) Helped to reduce post harvest losses. Increased efficiency of mineral usage by plants( this prevents early exhaustion of fertility of the soil) Enhanced nutritional value of food e.g., Vitamin A enriched rice.

1. 2. 3. 4. 5.

In addition to these uses, GM has been used to create tailor made plants, which could decrease the amount of pesticides used. Bt. toxin is produced by a bacterium called Bacillus thuringiensis (Bt. for short). Bt. toxin gene has been cloned from the bacteria and has been expresses in plants to provide resistance to insects without the need for insecticides; in effect created a bio-pesticide. Examples are Bt. cotton, Bt. Corn, rice, tomato, potato and Soya bean etc.)

19.

What are the agricultural applications of plant biotechnology?

During the last two decades plant protoplast, cell and tissue and organ culture have developed rapidly and became a major bio technological tool in agriculture, forestry and horticulture. M.S.Swaminathan emphasized the significance of biotechnological application of invitro cultural plant protoplasts/cell/ tissues as follows Tissue culture application in order to capitalize upon the totipotency of cells. Cell and protoplast cultures coupled with DNA vectors to overcome. Problems caused by barriers to gene transfer. Culture of plant cells for the production of useful compounds. Extension and increase of efficiency of biological nitrogen fixation.

Hybrid improvement: Development of cell fusion and hybridization techniques have solved the problems of incompatibility of plants and extended the scope of production of new varieties within a short span of time. Encapsulated (Artificial) seeds: These are somatic embryos covered with a protected gel. Gel in encapsulated seeds acts as a coat and artificial endosperm provides nutrient. Disease- resistant Plant Production: Vegetatively propagating plant species are mostly systematically infected by micro-organisms like bacteria and viruses, fungi and nematodes. In order to ensure highest possible yield and quality, it is essential to provide disease-free stock plants to growers. Tissue culture techniques have solved this problem and minimized the time of biological testing. Production of virus-free plants: Approximately 12% of viruses are transmitted through seeds which cause great loss. Tissue culture technique can be utilized for the production of virusfree plants either through meristem culture or selective chemotherapy of larger explants from donor plants or dormant propagules.

Invite selection of cell lines for disease resistance: Callus cultures are being widely used for study of expression of race-specific and non host resistance and offer sever advantages over suspension culture. Subclones regenerated from callus cultures have been found more resistant than the original material against eyespot( Helminthosporium sacchari), Dizi disease, decoxy mildew( sclerospora sacchari) Transfer of nif-Gene to Elukaryotes: Nitrogen fixing ability and genetic character exists in prokaryotic diatrophs. Researchers are still engrossed in solving this problem through tissue culture techniques coupled with the recombinant DNA technology or genetic engineering. Somoclonal variation: It is a term used to describe the variations in plants that have been produced by plant tissue culture which proved an alternative tool to plant breeding for generating new varieties that can exhibit disease resistance and improvement in quality and yield in plants such as oil seeds, fruit crops, legumes, tubur crops and cereals. 20. How radioactive biotechnology? pollutants are disposed/converted using

Radioactive waste containing heavy metals like mercury and toxic organic solvents like toluene are extremely costly to treat. Radiation resistant bacteria that can dispose of heavy metals are being developed to help clean up soil and water contaminated by radioactive waste. Deinococcus radiodurans is the most radiation-resistant organism known to man. This bacterium can grew at levels of radioactivity of 60 grays (Gy) per hour- about ten times the lethal dose for a human. Hassan Brim and colleagues at the University of the Health Sciences at Bethesda, Maryland, USA have genetically engineered this organism with a gene from E.Coli that can convert the highly toxic Hg(II) into less toxic elemental mercury and a gene from a soil bacterium, Pseudomonas Putida, that can degrade toluene. The end result is a strain of Deinococcus that converts mercury, breaks down toluene to use as a source of carbon and energy and does all this while thriving at radiation levels that no other organism can resist.

21. What is genetic engineering? Write about Genetic Engineering of plants. The technique of removing, modifying or adding genes to a DNA molecule (of an organism) in order to change the information it contains if Genetic Engineering. By changing this information, genetic engineering changes the type or amount of proteins an organism is capable of producing, thus enabling it to make new substances or perform new functions. The objective of genetic engineering is to produce desired characteristics and to eliminate undesirable ones. Examples of desired characteristics sought for plants include fast growth, pest resistance, drought resistance and stress resistance, larger and economically important parts. Organisms modified by genetic engineering are sometimes referred to as transgenic, Bioengineered, or GH organisms. Genetic engineering holds advantage over conventional plant breeding can only combine closely related plants. Genetic engineering permits the transfer of genes between organisms that are not normally able to cross breed because they are not genetically compatible. They can come from another plant species, or even from a completely different organism (e.g., bacterial genes). The diagrammatic explanation of process of genetic engineering: Genetic engineering of plants usually makes use of vector (carrier) which can transfer desirable segment of DNA to some plants. It is generally done by some bacteria or viruses (e.g. bacteriophages). When the bacterium infects the plant, it penetrates the plants cells and transfers its modified DNA to the plant. The DNA may also be introduced by a number of physical means once the DNA reaches the cell nucleus, it inserts itself at random into one of the host chromosomes and can express the desires character. The GH plant is then grown from the transformed cell. A number of economically valuable characteristics have been introduced into plants by genetic engineering. Most of the GH crop plants used so far have transgenes that provide resistance to herbicides or insects. The objective of genetic engineering is to improve crop production and soil management. Advance research is now exploring how to increase the variety of transgenic characteristics to include resistance to drought, heat, cold, acid soils and heavy metals. These characteristics are supposed to increase the range of soils and climates that are able to support agriculture.

22.

Define Genetic Engineering and what are its applications?

The manipulation of genetic make up of living cells by inserting desired genes through a DNA vector is called Genetic Engineering. Applications: Genetic engineering has many practical applications in agriculture, health and industry. Agriculture: Genetic engineering is used to produce 1. 2. 3. 4. 5. 6. Pest/ insect resistant plants. Oxygen tolerant plants. Petunia wit attractive flowers. Plants with high nutritional value. Nitrogen fixing plants. Transgenic animals with desired character like high milk yielding etc.

Health Genetic engineering is used in health science to produce 1. 2. 3. 4. 5. 6. 7. Human insulin in bacteria. Vaccines against cholera, typhoid, small pox etc Monoclonal antibodies to treat severe diseases. Interferons to treat viral diseases. Human growth harmone(Somatotropin) Diagnostic kits for genetic disorders and diseases. Genes to correct genetic disorders.

Industry In industries genetically engineered organisms are employed in the commercial production of many valuable goods. Human insulin, somatotropin, interferons, human serum albumin, human clotting factors VIII & IX epidermal growth factor(EGF), diagnostic kits, tissue plasminogen activator, hepatitis B Vaccine, foot and moth disease vaccine, antibodies and steroid harmones. 23. What is the methodology of genetic engineering?

Genetic engineering is a noval technique to create new strains of organisms with desired characters in short duration. The various steps involved are: 1. Preparation of desired gene.

2. 3. 4. 5. 6. 24.

Isolation of DNA Vector Construction of recombinant DNA. Introduction of recombinant DNA into the host cell Selection and multiplication of recombinant host cells. Expression of cloned gene. What are genetically engineered microorganisms and what are its uses?

the genetically manipulated bacteria and yeast are collectively called genetically engineered microorganisms (GEMOs). The important uses of GEMOs are : Health care products: 1. Insulin: It is a harmone secreted by the B-cells of the islets of langerhans os pancreas whose deficiency leads to diabetics in man. Elililly(USA) manufactures human insulin in the name of humulin. 2. Human Growth Harmone: (hGH) is secreted by the anterior lobe of the pituitary gland which is also known as Somatropin and its deficiency leads to dwarfism in man. The cDNA of Somatropin was introduced into E.Coli through the plasmid PBR 322 which in turn produces hGH. 3. Interferons : Interferons are antiviral proteins which protect the cells from the second viral infection and are produced by infected animal cells. The cDNAs of various interferons were engineered E.coli cult was produce interferons such as IFN-ALPHA, IFN-BETA, LeIF-D, IFN-alpha1 etc 4. Cytokines: These are small proteins that stimulate the migration of cells to the source of cytokines. They attract invading cells and destroy them. Some stimulate the immune system. e.g: Interleukin-2, CD 4 antigen and tissue necrosis factor (TNF)

5. Blood products: Genetic engineering is used to produce certain proteins that act on blood or bold producing cells. Such proteins are often called blood products. They may act as thrombolytics or blood clotting agents or erythropoietin. 6. Growth factors: These are small protein molecules that stimulate or inhibit cell proliferation. Genetically engineered bacteria have been developed to produce growth factors for human use. 7. Bioactive peptides: These include small proteins which change certain physiological functions of the body. They can be produced from genetically engineered bacteria. ExamplesCalcitonin (for treating osteoporosis) Relaxin (to facilitate child birth) Alpha- antitrypsin (for treating emphysema) Glucagon (for treating hypoglycaemia) 8. Vaccines: The genetically engineered microbes express the cloned gene and produce HBs antigen which is isolated, purified and used for vaccination against some severe diseases such as hepatitis, foot and mouth disease, etc. such vaccines are called sub unit vaccines. Sometimes genes for antigens for more than one pathogens are linked together and cloned a bacteria produce multiple antigen peptides (MAPs). The MAPs are used for vaccination against more than one disease. GEMos 1. Genetically Engineered Nitrogen Fixers: The nif genes of klebsiella pneumonic have been transferred to harmless soil bacteria such as E.Coli, klebsiella aerogenes, Erwinia herbicola, salmonella typhimurium etc. these genetically engineered microbes are capable of fixing the atmospheric nitrogen in the soil. They are used as bio fertilizers.

2. Genetically Engineered Bio control Agents: The soil bacterium Pseudomonos fluorescenes is antagonistic to many plant pathogens, but it does not produce endotoxins. Bt toxin gene of Bacillus thurugenisis is transferred to Pseudomonus fluroscene. The genetically engineered P.flourscence produces Bt. Toxins when it is sprayed on the crops, it kills crop pests ans controls plant pathogens. 3. Pollution control : Genetically engineered microbes are used to degrade pollutants in the environment. New strains of Pseudomonas are used to clear up oil spills and hydrocarbons. Candido tropicalis is genetically manipulated to degrade aldrin, DDT and dioxin. 25. What is Gene Therapy? Treatment of genetic disorders by introducing proper genes into cells of the target organ is called gene therapy. The gene used in gene therapy is often called gene drug. The drug gene may be introduced into somatic cells or germ cells or zygotes. If genes are introduced into somatic cells, it is called Somatic cell gene therapy and if introduced into eggs or zygotes, it is called germ line gene therapy The gene drug may be 1) The correct gene substitute for the defective gene. 2) A gene overproducing a protein whose deficiency causes the genetic disease. 3) DNA that produces antisense RNA that binds with mRNA of the defective gene and stops its expression. The gene drug may be introduced into target cells by using retrovirus, adenovirus, liposomes, transfection, biolistics or electroporation. The resulting recombinant cells are implanted into the target organ Gene therapy is used to cure asthma, melanoma etc. in man.

26.

What are the possible Dangers of GEOs (Genetically Engineered Organisms)?

It is feared that the GEOs will once be a source of threat to the mankind. The following are the possible dangers likely to be caused by GEOs. 1. Persistence of Antibiotic Resistance Gene: The antibiotic resistance gene of GEOs may once go to a pathogenic bacterium that causes diseases in man and domestic animals. Such diseases cannot be cured with antibiotics being used today. 2. Transfer of insect resistance from transgenic plants to wild relatives: The Bt gene of transgenic plant may get transferred to its wild relatives and weeds. If so, the pest population as a whole will be eliminated from the area. In ecological point of view, the pests too play some role in the environment for its sustainability. So the complete elimination of a species is likely. 3. Transfer of Insect resistance Gene from transgenic plants to Microbes: Bt. Gene of GEOs may go to the intestinal bacteria by transformation which causes illness to humans. 4. Development of more powerful pathogens: The GMOs may escape from the laboratory and exchange their genetic material with some pathogenic microbes making them more deadly. 5. Disease Epidemics: If transgenic plants are exploited continuously in a habitat, the other varieties will be left out. Single unique species in an area may result in severe disease epidemics and crop loss. 6. Disturbance of Ecological scenario: Native organisms may get disturbed and eliminated due to exploitation of GEOs. This is harmful to ecological balance of the habitats.

7. Plant products causing Allergy: The altered nutritive products of transgenic plants may cause allergy to man during consumption. 8. Side effects of healthcare products from GEOs are feared. 9. Inactivation of Genes: GMOs may exchange their altered genes with their parent genotypes and wild relatives, which will disturb the genetic make up of the plants. 10. Use of pathogens: GMOs made from wild pathogens may cause diseases in sensitive species. 27. What are the biohazards of rDNA technology? The bad effects of GEOs are known as biohazards of rDNA technology. Some of them which are confirmed experimentally are: 1. Super AIDS virus: Unfortunately, when a team of scientists in USA, injected HIV into the body of mice, it combined with another virus and became a more powerful virus named as super AIDS virus. It is highly dangerous one as it can be transmitted by air too. 2. Allergy reaction of Soyabean: The people allergic to Brazil-nut are also allergic to the protein of the transgenic soyabean. When a British company introduced some genes of Brazil-nut into soyabean, an allergic reaction ranged from mild itching to sudden death has taken place. 3. Side effect of Human Growth Harmone: It is proved that administration of overdose of hGH causes leukemia in children.

4. Eosinophilia-myalgia syndrome(EMS): Regular consumption of the amino acid tryptophan produced from genetically engineered E.Coli K 12 causes a disease called Eosinophiliamyalgia syndrome (EMS). 5. Cancer caused by Bovine Somatotropin ( bST): Bovine Somatotropin is manufactured from genetically E.Coli. The opponents of rDNA technology products have been advertising that the milk of the bST treated cows causes cancer and mastitis. 28. What is a Gene bank and what is its importance? A collection of bacterial clones each of which contains a unique DNA fragment of a source organism is called gene bank or gene library. It contains almost all DNA of the source organism. Each segment of the source DNA remains in the rDNA inside the bacterial cell. To establish gene banks, the individual fragments of source DNA are inserted into suitable vectors by shotgun cloning. If the DNA fragments are obtained from genomic DNA of a source organism, the library is called genomic gene library or gene library. As all DNA are maintained in bacterial clones, the library is also known as Clone Bank. If CDNA clones made from m RNAs of an organism are used to make the library, then it is said to be a cDNA library. The cDNA library has no intron sequence. Gene banks are usually 95-99% complete. Plasmids, M 13 based vectors, phage ** derived vectors and yeast artificial chromosomes (YAC) are used to construct gene banks. Human genomic DNA can be cloned in 5,00,000 bacterial clones. If ** phages are employed as gene cloning vectors, the same can be cloned on 2,00,000 bacterial clones through cosmid vectors. YAC carries relatively large DNA fragments. Hence only 10000 yeast cell clones are enough to hold all DNA of man. Importance of Gene Banks: 1. Gene banks provide information about the structure and organization of genes present in eukaryotic genome.

2. Some selected clones contain desirable genes and are mass cultured for the preparation of desired products from the culture. Johnson used this method in the preparation of human insulin from the bacteria E.Coli. 3. The construction of gene map of chromosomes is now possible by using this method. Ruddle et al used this method in constructing the gene map of a specific human chromosome. 4. Shortle et al used this technique for the mutational studies either by deletion or by the addition of base substitution to the individual gene. 5. It is an ideal method for preserving the genes in bacterial cells. 6. Puhler et al ( 1979) isolated different clones of bacteria which contain all the nif genes of a nif-gene cluster. Thus they discovered the total number of operons present in the nif- gene cluster. 7. Gene banks provide desired DNAs ( genes) for gene manipulation. 29. Enumerate the applications of plant tissue culture technique

The invitro culture of plant cells or tissues in an artificial medium is said to be plant tissue culture. Plant tissue culture has several applications in the field of agriculture, horticulture, forestry, pharmacology, medicine and environment. Some important applications are 1. It is possible to produce a large number of plants within a short time and space through tissue culture. Mass propagation of plants through tissue culture is known as Micro propagation. This technique is applied for propagation of ornamental plants, orchids and other exotic plants, fruits and plantation trees etc., 2. Plants regenerated through tissue culture show variations called somaclonal variations, which are used for crop improvement. 3. In some plants like Papaya, female plants are needed in greater propagation to produce more fruits per cultivated area. Female plants are selectively produced through tissue culture. 4. Virus diseases from vegetatively propagated plants can be prevented by producing virus free plants from shoot-tip cultures.

5. Seed dormancy problems can be solved by embryo culture, which also helps in production of rare hybrid plants. 6. Artificial or synthetic seeds are produced from somatic embryos by encapsulation with sodium alginate. These can be easily stored, germinated and transported. 7. Haploids are produced through another culture and fertile homozygous diploid plants are produced from them by doubling of chromosomes with colchicine. 8. Tissue culture of medicinal plants helps by the transfer of foreign genes is completely dependent on plant tissue culture. 9. The production of transgenic plants by the transfer of foreign genes is completely dependent on plant tissue culture. 10. Conservation of plant biodiversity is the need of the hour. Storage and preservation of germ plasm is possible through a cold storage procedure of plants as well as plant parts called Cryo preservation.

30.

Explain briefly the steps involved in the tissue culture.

The term plant tissue culture is normally used as a general term to include the cultivation of all plant parts whether a single cell, a group of cells or an organ on an artificial nutrient medium. The following are the steps involved in plant tissue culture. 1. Preparation of nutrient culture medium: The nutrient culture medium is a mixture of various essential nutrients containing all the required macro and micro nutrients, amino acids, vitamins and carbohydrates. These nutrients are mixed in distilled water and the PH adjusted to the required level (5.6 to 6.0). The medium is solidified by the addition of agar for providing support during culture of plant parts. The most popular medium is the murashige and skoog(MS) medium.

2. Sterilization of the culture medium: The culture medium is sterilized to kill the microorganisms. The sterilization is carried out in a steam sterilizer called the autoclave for 15 minutes at 1210 at 15 lb pressure. They are checked for growth of microorganisms the next day and only the non-contaminated ones are used for inoculation of seeds or explants and incubated. 3. Preparation of explants: Any plant part such as root, stem etc., used as inoculum to grow into full fledged plant or organs invitro is called explant. The explants are cleaned with a liquid detergent in running water and surface sterilized with disinfectant solutions like sodium hypochlorite and rinsed thoroughly with sterile ( autoclaved) distilled water. Seeds are surface sterilized with 0.1% mercuric chloride, rinsed in sterile distilled water and inoculated on MS based medium to obtain the aspetic seedlings invitro. 4. Inoculation of explants: The transfer of explants onto the sterile nutrient culture medium taken in culture vessels is called inoculation. The inoculation of explant is carried out in the laminar air-flow chamber, which maintains an aseptic environment. 5. Inocubation for growth: The cultures are incubated for 3 to 4 weeks during which, the cells of the explant absorbs the nutrients, grow and undergo repeated divisions to produce a proliferating undifferentiated mass of cells known as callus or produce shoots or roots directly ( organogenesis). Alternatively, the explant develops an embryogenic callus through embryogenesis from which embryoids are produced. The somatic embryos are transferred to other culture media for development into complete plants. These embryos can be encapsulated in sodium alginate for storage and transport. They are known as synthetic or artificial seeds. 6. Acclimatization of plantlets and transfer to pots: The plants are washed gently to remove the culture media and planted in plastic pots containing soil-rite. The pots are covered with polythene bags, maintained in the laboratory for 1 to 2 weeks and are gently removed

gradually for acclimatization. When the plant appears strong and healthy, it is transferred to a regular pot.