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BIOTECHNOLOGY AND ITS APPLICATION

INTRODUCTION 

Biotechnology is defined as the technological exploitation of microorganisms, plant cells, animal cells and biological processes (including genetic engineering) for obtaining useful materials for agriculture, industry, domestic, medical and other uses.

BIOTECHNOLOGICAL APPLICATIONS IN AGRICULTURE

  • Improvement in the agricultural production is the most urgent need of the time in the context of rapid increase of population.
  • Agrochemical based agriculture, organic agriculture and genetically engineered crop based agriculture will increase food production.

GENETICALLY MODIFIED ORGANISMS

  • Plants, bacteria, fungi and animals whose genes have been altered by manipulation are called genetically modified organisms.
  • A transgenic crop is a crop that contains and expresses a transgene. 
  • It has two unique advantagesFirstly any gene (from any organism or a gene synthesized chemically) can be used for transfer, and secondly the change in genotype can be precisely controlled since only the transgene is added into the crop genome. For e.g., Hirudin is a protein that prevents blood clotting. The gene encoding hirudin was chemically synthesized and transferred into Brassica napus, where hirudin accumulates in seeds. The hirudin is purified and used in medicine.
  • The GM crops are fast becoming a part of agriculture throughout the world because of their contribution to the increased crop productivity and to global food, feed and fiber security, besides their use in health-care and industry. However, the constraints associated with public acceptance of transgenic crops continue to be important challenges facing the global community. 
  • The following are the major concerns about GM crops and GM food : 
    • The safety of GM food for human and animal consumption (e.g., GM food may cause allergies).
    • The effect of GM crops on biodiversity and environment.
    • The effect of GM crops on non-target and beneficial insects/microbes.
    • Transgenes may escape through pollen to related plant species (gene pollution) and may lead to the development of super weeds.
    • The GM crops may change the fundamental nature of plants as the genes from animals (e.g., fish or mouse) are being introduced into crop plants.
    • The antibiotic resistance marker genes used to produce transgenic crops may horizontally transfer into microbes and thus, cause problem of antibiotic resistance in human and animal pathogens (i.e. transgenes may move from plants to gut microflora of humans and animals).
    • The GM crops may lead to the change in the evolutionary pattern.
  • GM crops are already in cultivation in U.S.A., Europe and several other countries. 

Examples
Bt Cotton, tomato variety, etc.

BT COTTON
  • Bt means Bacillus thuringiensis. It is a gram positive dwelling bacterium. It produces crystal (Cry) proteins. This Cry protein is toxic to larvae of certain insects. Each Cry protein is toxic to a different group of insects. The gene encoding cry protein is called cry gene. This Cry protein  is isolated and transferred into several crops. 
A crop expressing a cry gene is usually resistant to the group of insects for which the concerned Cry protein is toxic. There are a number of them, for example, the proteins encoded by the genes cryIAc and cryIAb control the cotton bollworms, that of cryIAb controls corn borer.
  • Some strains of Bacillus thuringiensis produce proteins that kill certain insects such as lepidopterans (tobacco budworm, armyworm), coleopterans (beetles) and dipterans (flies, mosquitoes). B. thuringiensis forms protein crystals during a particular phase of their growth. These crystals contain a toxic insecticidal protein. The Bt toxin protein exists as inactive protoxins but once an insect ingest the inactive toxin, it is converted into an active form of toxin due to the alkaline pH of the gut which solubilise the crystals. The activated toxin binds to the surface of midgut epithelial cells and create pores that cause cell swelling and lysis and eventually cause the death of the insect.
  • It is a genetically engineered cotton to control tobacco budworms, bollworms and pink bollworms.

TRANSGENIC VARIETY OF TOMATO
Flavr Savr is formed due to the inhibition of polygalacturonase enzyme which degrades pectin, so that tomato variety remains fresh and retain flavour much longer.

TRANSGENIC PLANT OF TOBACCO
A nematode Meloidogyne incognita infects the roots of tobacco plants and causes a great reduction in yield. Using Agrobacterium vectors, nematode specific genes were introduced into the tobacco plant. The introduction of DNA produced both sense and antisense RNA in the host cells. These two RNA's being complementary to each other formed a double strand that initiated RNAi (= RNA interference), a method of cellular defence that takes place in all eukaryotic organisms) and thus, silenced the specific mRNA of the nematode. The result was that the parasite could not survive in a transgenic host expressing specific interfering RNA. The transgenic plant thus, got itself protected from the parasite.

APPLICATIONS OF TRANSGENIC PLANTS
  • They have proved to be extremely valuable tools in studies on plant molecular biology, regulation of gene action, identification of regulatory promoter sequences etc.
  • Specific genes have been transferred into plants to improve their agronomic and other features.
  • Genes for resistance to various biotic stresses have been engineered to generate transgenic plants resistant to insects, viruses, etc.
  • Several gene transfers have been aimed at improving the product quality, e.g., protein or lipid quality etc. of transgenic plants.
  • Transgenic plants are aimed to produce novel biochemicals like interferon, insulin, immunoglobulins, etc.
  • Transgenic plants have been produced that express a gene encoding an antigenic protein from a pathogen. 
  • They made crops which are more tolerant to abiotic stresses (cold, drought, salt, heat).
  • They have 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 soil).
  • Enhanced nutritional value of food, e.g., Vitamin A enriched rice.

PROBLEMS RELATED TO GM FOODS
  • The transgene product may produce toxicity or allergies. 
  • The enzyme produced by the antibiotic resistance gene could cause allergies, since it is a foreign protein. 
  • The bacteria present in the alimentary canal of humans could take up the antibiotic resistance gene that is present in the GM food.

NOTES
  • Transgenic plant was obtained through recombinant DNA technology. First transgenic plant was tobacco. It contains resistant gene against weedicide (Glycophosphate).
  • First transgenic animal was mouse containing gene for growth hormone.
  • First introduced transgenic crop in India (2002) is Bt-cotton.
  • In pollution control, microbes have been engineered to break up the crude oil spills.
  • Dr. Ananda Mohan Chakraborthi introduced plasmid from different strains in to single cell of Pseudomonas putida. The result was new genetically engineered bacterium which would clean oil spills called “Superbug” (oil eating bug.)

Table : Transgenic plants

BIOTECHNOLOGICAL APPLICATION IN MEDICINES

  • The recombinant DNA technological processes have made vast impact in the field of health and medicine by producing safe and more effective therapeutic drugs.
  • These drugs do not induce immunological reactions as some other drugs obtained from non human sources (e.g., insulin from animals).

GENETICALLY ENGINEERED INSULIN

  • Insulin is a pancreatic hormone essential for maintenance of glucose-glycogen balance in the body. In its deficiency, the glucose content of blood rises while the cells fail to take up glucose from blood. It results in diabetes mellitus. The disease is overcome by injection of small doses of insulin.
  • Human insulin (humulin) was first genetically engineered product produced by an American firm Eli-lilly (5th July 1983). Insulin was earlier extracted from the pancreas of slaughtered cattle and pigs. Such insulin though caused some patients to develop allergy and other types of reactions to the foreign proteins. Insulin consists of two short polypeptide chains A and B which are linked together by disulphide bridges. In mammals, insulin is synthesized as a prohormone, which contains an extra stretch called C-peptide, which is not present in the mature insulin.
Fig. : Maturation of pro-insulin into insulin (simplified)

GENE THERAPY

  • It is a therapeutic treatment of defective heredity by introduction of normal healthy and functional genes which also silence the defective genes of an individual.
  • The first clinical gene therapy was given to a 4 year old girl in 1990 suffering with SCID (Severe Combined Immuno Deficiency) with adenosine deaminase (ADA) deficiency. ADA deficiency is a very rare genetic disease. This enzyme is important for the immune system to function.
  • It is caused due to deletion of the gene for adenosine deaminase and treated by gene therapy.
  • About 25% of infants with SCID disorder lack the enzyme adenosine deaminase (ADA).
  • ADA enzyme is involved in purine metabolism.
  • These patients have no functioning T & B lymphocytes.
  • The affected child of SCID develops recurrent infection early in life because they have no immune response against invading pathogens.
  • The ideal approach would be to give the patient a functioning ADA by gene therapy.
  • Success has been achieved in few other cases also like cystic fibrosis, atherosclerosis. Work on gene therapy for thalassemia, muscular dystrophy and several other disorders is in progress.
  • Major requirement of gene therapy is the isolation of the desired gene, its amplification in a retrovirus for removal of viral genes and insertion of a desired gene and a helper retrovirus lacking packing sequence.

VACCINE PRODUCTION

  • Vaccine is a liquid containing dead, attenuated form or antigen of a pathogen which can be infected or taken orally to provide immunity towards that pathogen.
  • Vaccines which are produced by genetic engineering are for hepatitis-B and herpes virus.

Table : Applications of Recombinant DNA products

Table : Application of Genetically Engineered Microbes
 

MOLECULAR DIAGNOSIS

  • Recombinant DNA technology is one of the important tools for the diagnosis of several diseases. 
  • The diagnostic technique involves the construction of probes consisting of short segments of single stranded DNA attached to a radioactive or fluorescent marker. Such probes are used to identify infectious agents such as Salmonella (cause food poisoning), Staphylococcus (pus forming bacterium), hepatitis a virus, HIV, muscular dystrophy, cystic fibrosis and so on. 
  • Recombinant DNA technology can also be employed to predict the inheritance of genetic disorders from carrier parents. The chances of birth of an affected child can be predicted by testing DNA of repetitive prospective genetic disorder carrier parents.
  • Some of the techniques that are included in molecular diagnosis are recombinant DNA technology, polymerase chain reaction (PCR), Enzyme linked immuno sorbant assay (ELISA). They serve the purpose of early diagnosis.

TRANSGENIC ANIMALS

  • Transgenic animals are those animals which have specific foreign genes. They have their DNA manipulated to possess and express an extra (foreign) gene.
  • Foreign DNA is introduced into the animal using recombinant DNA technology.
  • Transgenic animals are those animals that have their DNA manipulated to possess and express an extra (foreign) gene. 
    • Normal physiology and development : Transgenic animals can be exclusively designed to allow the study of how genes are regulated, and affect the normal functions of the body and its development. E.g., study of complex factors involved in growth such as insulin-like growth factor. 
    • Study of disease : Transgenic animals are designed to increase the understanding of the process through which genes contribute in the development of disease. These are specially 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 alzheimer’s.
    • Biological products : 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 (α-1antitrypsin) used to treat emphysema. Similar method is required for the treatment of phenylketonuria (PKU) and cystic fibrosis. In 1997, the first transgenic cow, Rosie, produced human protein-enriched milk (2.4 grams per litre). The milk contained the human alpha-lactalbumin and was nutritionally a more balanced product for human babies than natural cowmilk.
    • Vaccine safety : Transgenic mice are being developed for use in testing the safety of vaccines before they are used on humans. Transgenic mice are being used to test the safety of the polio vaccine. 
    • Chemical safety testing : This is known as toxicity/safety testing. The procedure is the same as that used for testing toxicity of drugs. Transgenic animals are made to carry genes which make them more sensitive to toxic substances than non-transgenic animals. They are then exposed to toxic substances and the effects are studied. Toxicity testing in such animals will allow us to obtain results in less time.
    • Developing animals specially created for use in xenografting.

Table : Transgenic animals

ETHICAL ISSUE

ETHICS

  • Bioethics includes a set of standards by which a community regulates its behaviour and decides as to which activity is legitimate and which is not. Therefore, bioethics may be viewed as a set of standards that may be issued to regulate our activities in relation to the biological world.
  • The Indian Government has set up organisations such as GEAC (Genetic Engineering Approval Committee), which will make decisions regarding the validity of GM research and the safety of introducing GM organisms for public services.
  • The major bioethical concerns pertaining to biotechnology are summarised below.
    • Use of animals in biotechnology causes great suffering to them.
    • When animals are used for production of pharmaceutical proteins, they are virtually reduced to the status of a 'factory'.
    • Introduction of a transgene from one species into another species violates the ‘integrity of species’
    • Transfer of human genes into animals (and vice-versa) dilutes the concept of ‘humanness’.
    • Biotechnology is disrespectful to living beings, and exploits them for the benefit of human beings.
    • Biotechnology may pose unforeseen risks to the environment, including risk to biodiversity.

BIOSAFETY ISSUE

BIOPIRACY

  • Many organisations and multinational companies exploit or patent biological resources or bioresources of other nations without proper authorisation from the countries concerned is known as biopiracy. 
  • The industrialised nations are rich in technology and financial resources but poor in biodiversity and traditional knowledge related to the utilisation of the bioresources. In contrast, developing nations are poor in technology and financial resources, but are rich in biodiversity and traditional knowledge related to bioresources.
  • Biological resources or bioresources include all those organisms that can be used to derive commercial benefits. Traditional knowledge related to bioresources is the knowledge developed by various communities over long periods of history, regarding the utilisation of bioresources, e.g., use of herbs, etc. as drugs.
  • Institutions and companies of industrialised nations are collecting and exploiting the bioresources, as follows:
    • They are collecting and patenting the genetic resources themselves. For e.g., a patent granted in U.S.A. covers the entire 'basmati' rice germplasm indigenous to our country.
    • The bioresources are being analysed for identification of valuable biomolecules. A biomolecule is a compound produced by a living organism. The biomolecules are then patented and used for commercial activities.
    • Useful genes are isolated from the bioresources and patented. These genes are then used to generate commercial products.
A west African plant Pentadiplandra brazzeana produces a protein called brazzein which is approximately 2000 times sweeter than sugar. It is proposed to transfer the brazzein gene into maize and express it in maize kernels.
But the protein brazzein was patented in U.S.A. subsequently, the gene encoding brazzein was isolated and patented in U.S.A.

BIOPATENT

  • A patent is a right granted by a government to an inventor to prevent others from commercial use of his invention. 
A patent is granted for
    • an invention (including product).
    • an improvement in an earlier invention.
    • the process of generating products, and
    • a concept or design.
These products are called biopatents because they are granted for biological entities.
  • Biopatents are awarded for the following :
    • Strains of microorganisms. 
    • Cell lines 
    • Genetically modified strains of plants and animals. 
    • DNA sequences. 
    • The proteins encoded by DNA sequences. 
    • Various biotechnological procedures. 
    • Production processes. 
    • Products
    • Product applications.
E.g., one patent covers ‘all transgenic plants of Brassica family’. Such broad patents are considered morally unacceptable and fundamentally inequitable.

BIOWAR

  • Biowar or biological war is the use of biological weapons against humans or their crops and animals. 
Bioweapons are devices that carry and deliver to the target organism in the form of pathological biological agents or toxins. 
Bioweapon agent is kept in a suitable container so that it remains active and virulent during delivery. This container could be delivered in various ways, including missiles and aircraft. The use of biological agents in war may date back to the 5th century B. C.

  • Some of the potential pathogen for bioweapons are those that causes anthrax, smallpox and botulinum toxin. These bio weapons are sent either through letters (envelopes) as a powder or in the form of spray.
  • The possible defences against bioweapons include the use of respirator or gas mask, vaccination, administration of appropriate antibiotics, and decontamination. In addition, sensitive detection systems should be developed to control and minimise damage.
  • Benefits of Biological Weapons over Conventional weapons:
    • Biological weapons or agents for biological warfare are not costly and can be produced with ease. 
    • Detection is almost impossible.
    • Use is likely to increase.
    • They are more dangerous than conventional weapons.
  • Potential Biological Weapon Agents are:
    • Anthrax → Bacillus anthracis.
    • Botulinum toxin (Botulin) → Clostridium botulinum. 
    • Plague → Yersinia pestis.
    • Viral encephalitides → Alpha virus.

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