What is Biotechnology?

Biotechnology is the use of living organisms and their internal systems for the benefit of
the human being. With the advancement of various molecular techniques, scientists
have used biotechnology in various fields, e.g. for the production of genetically modified
(GM) crop, new drugs, vaccines, enzymes, chemicals for industrial use, diagnosis, to
increase the yield of the crop, waste management, etc.

Applications of Biotechnology
Biotechnology is widely used in different fields to modify and produce useful products
for human benefit. These applications include:

Applications of Biotechnology in Agriculture

The application of Biotechnology in the field of agriculture is known as Green
Biotechnology or Green Revolution. Biotechnology has contributed a lot to the field of
agriculture, and they include:

1. Organic agriculture
2. Agro-chemical based agriculture
3. Genetically engineered crop-based agriculture

The application of biotechnology has tripled the yield and the food supply. Apart from
regular crop production, pest-resistant plants and genetically modified crops are
introduced to increase food production and to feed the growing human population.

Genetically modified crops have their genes altered by inserting the genes with desired
characteristics. Bt cotton, Bt brinjal, and golden rice are a few examples of Genetically
modified crops.

Applications of Biotechnology in Medicine

The recombinant DNA technology has added to the advancements in the field of
healthcare by permitting mass production of safe and more effective therapeutic drugs.

The drugs are mainly produced by the process of genetic engineering. Humulin, the
genetically modified insulin that is used to treat diabetes, is produced through genetic
engineering.
Biotechnology has developed gene therapy which helps in the removal of genetic
disorders in the embryo. Other applications of biotechnology in the field of medicine and
molecular diagnosis include:

1. ELISA
2. PCR

Transgenic Animals
Transgenic animals can be defined as those animals in which a new or altered gene
has been experimentally inserted into the genome by genetic engineering techniques.

A few examples of transgenic animals are rats, rabbits, pigs, sheep, cows, fish, etc.
Among all other transgenic animals, the mouse is the existing transgenic animal.

The main aim behind the creation of transgenic animals are:

1. For the production of biological products.

2. To study the different types of diseases.
3. To study the contribution of genes in the development of the disease.
4. For testing the safety of vaccines and toxicity of drugs before they are used on humans.
5. To study how genes are regulated and how they affect the normal functioning of the body
and its development.

Applications in Aquaculture
Biotechnology applications help in the improvement of the quality and quantity of fish.
The gonadotropin-releasing hormone is introduced into the fish to enhance breeding.
This helps in enhancing growth and improving their genetic characteristics. It also
prevents a number of diseases.

Production of Antibiotics
Biotechnology helps in the production of vaccines, antibiotics and artificial hormones
using plants. Genes with desired characteristics are introduced into the plants to
manufacture the encoded proteins. Edible vaccines are cost-effective, and can be easily
stored and administered in the body. These are used to cure diseases such as measles,
hepatitis, cholera, etc.

Applications of Biotechnology
Earlier the use of biotechnology was limited to making bread, alcoholic beverages and
cross breed animals and crops. Modern biotechnology makes use of the advancement
in science such as genetic engineering and recombinant DNA technology with great
effect and has expanded the application of biotechnology for the benefit and well being
of humans.

1. Application of Biotechnology in Agriculture

The “Green revolution” used the application of pesticides and fertilizers to increase the
production of crops. This resulted in excessive use of chemicals, thereby polluting the
soil and environment as well as various harmful effects to the consumer.

Application of biotechnology in the field of agriculture has greatly reduced the usage of
fertilizers and chemicals.

Genetically Engineered Crops:

Using genetic engineering and recombinant DNA technology, genes for the desired trait
are introduced in the species. This type of genetically modified plant species is known
as GM crop.

These GM plants have many benefits:

• Crops can be made insect and pest resistant

• Nutritional value of the food can be increased
• Increases the yield by many folds
• Reduces the use of harmful chemicals and fertilizers
• Crops can be made more tolerant to environmental stress
• Efficient mineral utilization by plants
• Yield can also be increased by having a short maturing period

Examples of GM Plants

• Golden rice: Vitamin A enriched variety of rice.

• Bt. cotton, Bt. corn: Bacillus thuringiensis produces an insecticidal protein, known as Bt
toxin. These insect-specific Bt toxin genes are incorporated into the genome of certain plants
like cotton, corn, etc. These modified crops have insect resistant quality.
• Pest resistant tobacco plant: A nematode Meloidogyne incognita lives as a parasite in the
root of tobacco plants. The plant is made resistant to infection by using the RNA interference
technique. The nematode specific genes are introduced in the plant
using Agrobacterium vector. The transgenic plant produces double-stranded RNA, which
silences mRNA of the nematode and they are unable to survive.
 • Virus resistant plants: The gene encoding for the protein of virus coat is transferred to the
host plant resulting in the virus-resistant plant species. This has been used to produce virus-
resistant squash.
• Flavr Savr Tomato: Gene responsible for the production of the enzyme polygalacturonase
is blocked. This enzyme is responsible for degrading pectin and softening of the fruit. The
transgenic variety of tomato can stay fresh and retain flavour for a longer duration.

2. Application of Biotechnology in Medicine

Biotechnology has vast applications in the medical field. Biotechnology is used for
therapeutic, diagnostic, scientific investigations for forensic studies, production of
vaccines, antibiotics and various drugs.

• Production of antibiotics, vaccines, enzymes and proteins: Using recombinant DNA

technology, many safe and therapeutic drugs have been produced. These drugs do not
induce an allergic reaction, which may be the case if the same product is isolated from any
animal source.Production of antibiotics and vaccines: Antibiotics are produced using
plants. Desired genes are incorporated in plants and targeted proteins are produced. Edible
vaccines have already been manufactured for some diseases, e.g. hepatitis B, measles,
cholera.Genetically Engineered Insulin: Insulin is used for the treatment of diabetes.
Earlier insulin was extracted from the pancreas of cattle and pigs, which has shown to induce
allergic reactions. Using recombinant DNA technology, genes coding for human insulin were
incorporated in the plasmid of non-pathogenic strains of E. coli. Forming chains of insulin,
which are extracted and combined by disulfide bonds. The recombinant human insulin is
known as Humulin. It is widely used to treat diabetic patients.Digestive
enzymes: Microorganisms can be modified to produce digestive enzymes. These
microorganisms can be colonised in the digestive tract to suffice for the insufficient
enzymes.Hirudin Protein: The gene coding for hirudin protein, which prevents blood clotting
is transferred into Brassica napus. The protein gets accumulated in the seeds, which can be
purified and used medicinally.Single Cell Protein (SCP): It is a microbial protein, which has
high-quality protein and less fat. It is used as a protein-rich supplement for the human diet.
• Gene Therapy: Gene therapy is used in correcting genetic defects in embryo and child.
Normal genes are delivered to the embryo or an individual with a faulty gene so that the
function of the faulty gene is restored. For the first time, gene therapy was performed on a
girl with adenosine deaminase (ADA) deficiency in 1990. This is caused due to the
deletion of the gene coding for ADA. Functional adenosine deaminase cDNA is incorporated
into the lymphocytes extracted from the blood of the patient using a retroviral vector and
reintroduced in the body. Genetically engineered lymphocytes have to be infused
periodically. The disease can be cured permanently If the gene is isolated from marrow cell
and it is introduced in the embryonic stage.
• Molecular Diagnostics: Biotechnology has proven useful in early diagnosis of
disease.PCR (Polymerase chain reaction): very low concentration of virus and bacteria can
be detected by amplifying the DNA. PCR is used to detect the HIV virus in AIDS patients.
Mutation of the gene in a cancer patient can also be detected by using this
technique.Elisa (enzyme-linked immunosorbent assay): Presence of antibodies synthesised
as a result of reaction with the antigen of the pathogen can be detected by this technique.
• Transgenic Animals: Transgenic animals are those animals whose genes are manipulated
to express a foreign gene. These transgenic animals are useful in many ways, e.g.
• Study gene regulation during normal growth and development
 • Understand the progression of the disease. Various treatments can be tried on
transgenic models with life-threatening diseases
• For the production of various biological products, e.g. gene coding for protein 𝛼-1-
antitrypsin used to treat emphysema was incorporated to form the transgenic animal
• Rosie, the first transgenic cow, produced milk containing a human protein, 𝛼-
lactalbumin
• Safety of vaccines can be tested on transgenic animals, e.g. the polio vaccine was
tested on transgenic mice
• Toxicity of drugs can be tested on transgenic animals

3. Application of Biotechnology in Industry:

Industrial processes can be made more efficient and environment-friendly by using
biotechnology.

• Using biotechnology enzymes can be produced in large quantities. This can be used as a
biocatalyst for the synthesis of many chemicals
• Genetically modified plants supply different resources like starch, fuel, pharmaceuticals in
huge quantity
• Proteases are used in detergents
• Biofertilizers production
• The processed food industry is based on biotechnology and the use of microorganisms for
the fermentation process
• Large scale production of fructose from glucose derived from inexpensive sources
• Corn can be used in place of petroleum to synthesize chemicals
• Biotechnology is useful in getting a better quality of products, e.g. cotton produced through
biotechnology is warmer, stronger and wrinkle resistant
• Production of biopesticide. Biopesticides act selectively by killing pathogens such as
bacteria, viruses and insects. These are not harmful like chemical pesticides
• Production of aquatic species by using biotechnological tools has greatly enhanced the
quality and growth of aquatic organisms

4. Environmental Application of Biotechnology

Biotechnology is useful in waste management and controlling pollutants and energy
production.

• Waste Management: Industrial and food waste are pretreated in enzyme bioreactors and
subsequently removed through the sewage system. Waste can be converted to bio-fuel.
Plant and vegetable materials can be degraded by microorganisms to make biodegradable
plastics.
• Bioremediation: Microorganisms are deliberately introduced to water, soil, etc. to degrade
the target pollutants. Bioprocessing reduces pollution by reducing harmful gases produced
due to combustion of coal and oil.
 • Energy Production: Bio-fuels and petroleum products are produced using biotechnology.
Bio-methane, Bio-diesel and ethanol are produced on a commercial scale. Biotechnology is
used to refine petroleum and coal from undesirable impurities. It makes oil less expensive
and easier to refine.

D
Difference between Electroporation and Microinjection
Foreign DNA is transmitted to host cells through a process called transformation.
Transformation results in the modification of the genetic composition of the organism.
There are numerous physical, chemical and biological transformation techniques.

There are direct and indirect approaches, respectively. Microinjection and

electroporation are two physical techniques that involve direct transformation. DNA is
incorporated into host cells through the process of electroporation, which creates small
pores in the living cell membranes. While a micropipette or a fine-tipped glass needle is
used in microinjection to deliver the DNA directly.

Electroporation
DNA is introduced into protoplasts and plant cells via the transformation procedure
known as electroporation. This method makes use of an electric pulse with high voltage.
Plant samples are incubated in a DNA-containing buffer solution. Then an electric pulse
with a high voltage is applied to the solution. Plant cell membranes develop high
voltage-driven pores that allow DNA to go within the cells and combine with the
genomic DNA of plants. The plants used and the conditions under which they are
treated determine how effective this procedure is.

Only 40-50% of cells obtain DNA when the transformation is performed using
electroporation. Additionally, this approach only allows 50% of the transformed cells to
survive. However, this procedure is simple to perform and does not modify the biological
configuration or function of cells. It can also be applied to various cells.

Microinjection
The transformation method of microinjection is particularly efficient when inserting DNA
into giant cells. The microinjection technique introduces DNA into animal cells (eggs,
oocytes, and embryos) or plant protoplasts using a micropipette (fine-tipped glass
needle). This technique is more appropriate for producing transgenic mice. This process
involves incorporating DNA straight into the cytoplasm or nucleus.

Microinjection is a method of direct transformation, much like electroporation.

Microinjection is carried out using a specialised microscope setup. The effectiveness of
this technique has been improved by computerised control of the microscope stage,
needle, holding pipette, and video technologies. A dye can be utilised to determine
transformed cells after DNA injection.

This technique is very trustworthy and effective. However, this procedure is expensive,
time-consuming, and labour-intensive. Furthermore, this technique can only be used to
treat a few cells.

Difference between Electroporation Method and

Microinjection Method
Electroporation Microinjection

t is a physical and direct transformation It is a physical and direct transformation

method which utilises an electric field to approach that inserts DNA into host cells
induce DNA into host cells. using a fine-tipped glass needle or
micropipette.
Can be used for a large number of cells Can be used to treat a small number of cells
Not time-consuming Time-consuming method
Induces microscopic pores in plasma Does not induce any microscopic pore in
membranes plasma membranes
Mainly used for protoplasts and plant cells Mainly used for animal cells

Does not require any specialised microscope Requires specialised and computerised
setup microscope setup

Selection of transformed cells is not very Selection of transformed cells is easy

easy

Thus, this is the main difference between microinjection and electroporation. However, both
procedures directly insert exogenous DNA into the host cells.

Q1
Why is electroporation more efficient than chemical transformation?
In general, electroporation offers higher transformation efficiencies and is less labour-intensive than
chemical transformation.

Q2

What is the usage of electroporation?

Electroporation, also known as electropermeabilisation, is a microbiological procedure in which cells are
exposed to an electrical field to enhance the cell membrane’s permeability and permit the introduction
of drugs, chemicals, or DNA.

Q3

What is the microinjection method?

Microinjection is the process of injecting a liquid element at a microscopic scale using a glass
micropipette. The target is usually a live cell, however, it may also be intercellular space.

Biotechnological Applications in Agriculture and Medicine:

1. Biotechnology essentially deals with industrial scale production of biopharmaceuticals and
biologicals. The applications of biotechnology include therapeutics, diagnostics, genetically
modified crops for agriculture, processed food, bioremediation, waste treatment and energy
production.
2. Biotechnology have the following three critical research areas:
(i) To provide the best catalyst in the form of improved organism, usually a microbe or pure
enzyme.
(ii) To create optimal conditions through engineering for a catalyst to act.
(iii) Downstream processing technologies to purify the protein/organic compound.
3. Biotechnological Applications in Agriculture
(i) Biotechnology applications in agriculture involve following three options:
(a) Agrochemical based agriculture.
(b) Organic agriculture.
(c) Genetically engineered crop-based agriculture.
(ii) Green revolution increased food production due to the use of:
(a) Improved crop varieties.
(b) Agrochemicals (fertilisers and pesticides).
(c) Better management practices.
(iii) Genetically Modified Organisms (GMOs) are plants, animals, bacteria and fungi whose
genes have been altered by manipulation.
(iv) Genetic modification in plants have lead to following:
(a) Crops became more tolerant to abiotic stresses, such as cold, drought, salt, heat, etc.
(b) Dependence on chemical pesticides reduced, i.e. pest resistant crops.
(c) Post harvest losses reduced.
(d) Efficiency of mineral usage increased in plants (preventing loss of soil fertility).
(e) Nutritional value of food is enhanced, e.g. vitamin-A enriched rice.
(f) Tailor made plants are created by using GM plants to supply alternative resources to
industries, in the form of starches, fuels and pharmaceuticals.
(v) Some of the applications of biotechnology in agriculture are the production of pest resistant
plants, which decrease the amount of pesticide used.
Bt toxin is produced by a bacterium and expressed in plants to provide resistance to insects, in
effect created a biopesticide, e.g. Bt cotton, Bt corn, golden rice, tomato, potato and soybean,
etc.
(a) Bt cotton is created by using some strains of a bacterium, Bacillus thuringiensis (Bt is short
form,).
(b) This bacterium produces proteins that kill certain insects such as lepidopterans (tobacco,
budworm and armyworm), coleopterans (beetles) and dipterans (flies and mosquitoes).
(c) B. thuringiensis forms protein crystals during a particular phase of their growth. These
crystals contain a toxic insecticidal protein.
(d) Bt toxin protein exist as inactive protoxins, but once an insect ingests the inactive toxin, it is
converted into an active form of toxin due to the alkaline pH of the gut, which solublise the
crystals.
(e) The activated toxin binds to the surface of midgut epithelial cells and create pores that cause
cell swelling and lysis leading to death of an insect.
(f) Specific Bt toxin genes were isolated from Bacillus thuringiensis and incorporated into
several crop plants as cotton.
(g) Most Bt toxins are insect-group specific. The toxin is coded by a gene named cry, e.g. the
proteins encoded by the genes cry IAc and cry IIAb control the cotton bollworms and cry IAb
controls corn borer.
(vi) Pest resistant plants are developed by using biotechnological processes.
(a) A nematode Meloidogyne incognita infects the roots of tobacco plants, which reduces the
production of tobacco.
(b) RNA interference (RNAi) process is used for cellular defence. It involves silencing of a
specific mRNA due to a complementary dsRNA. It occurs in all eukaryotic organisms as a
method of cellular defense.
(c) dsRNA binds and prevents translation of the mRNA (silencing).
(d) The source of this complementary RNA could be from an infection by viruses having RNA
genomes or mobile genetic elements (transposons) that replicate via an RNA intermediate.
(e) Agrobacterium vectors are used to introduce nematode-specific genes into the host plant. It
produces both sense and anti-sense RNA in the host cells.
(f) These two RNAs are complementary to each other and forms a double stranded RNA
(dsRNA) that initiate RNAi and hence, silence the specific mRNA of the nematode.
(g) The parasite cannot survive in transgenic host, expressing specific interfering RNA. The
transgenic plant thus, gets itself protected from the parasite.
4. Biotechnological applications in medicine have made immense impact in the area of
healthcare by enabling the mass production of safe and more effective therapeutic drugs.
(a) The recombinant therapeutics do not induce unwanted immunological responses
as in case of similar products isolated from non-human sources.
(b) Currently, about 30 recombinant therapeutics have been approved for human
use over the world. In India, 12 of these are presently being marketed.
I. Genetically engineered insulin leads to sufficient availability of insulin for the
 management of adult-onset diabetes.
(a) Insulin used for diabetes was earlier extracted from the pancreas of
slaughtered cattle and pigs. This caused allergy or other reactions in some
patients.
(b) Insulin consists of two short polypeptide chains, i.e. chain-A and B, linked
together by disulphide bridges.
Maturation of proinsulin into insulin (simplified)
(c) In mammals, insulin is synthesised as a prohormone (needs to be processed
before it becomes a fully mature and functional hormone) which contains an
extra stretch called the C-peptide.

(d) C-peptide is not present in the mature insulin $nd is removed during maturationinto insulin.
Thus, the main challenge for the production of insulin using rDNA techniques was getting
insulin assembled into a mature form.
(e) Eli Lilly an American company in 1983, prepared two DNA sequences corresponding to A
and B-chains of human insulin and introduced them in plasmids of E. coli to produce insulin
chains. Chains-A and B were produced separately, extracted and combined by creating
disulphide bonds to form human insulin.
II. Production of vaccines through genetic engineering such vaccines are called recombinant
vaccines also called ‘subunit vaccines’ or ‘second generation vaccines’, e.g hepatitis-B. These
are of two types:
(a) Protein vaccines use of specific protein produced by rDNA in vaccine.
(b) DNA vaccines use of genetically engineered DNA to be injected as vaccine to produce an
immunological response.
Hepatitis vaccine contains the viral envelope protein, hepatitis-B surface antigen (HB8 Ag). This
gene is isolated from yeast vectors.
Some protein coding genes isolated from pathogens are also incorporated and expressed in plants
produce antigens and are also called edible vaccines.
III. Gene therapy is a collection of methods that allows correction of gene defects, diagnosed in
a child or embryo.
(a) Genes are inserted into a person’s cells and tissues to treat a disease.
(b) Correction of a genetic defect involves delivery of a normal gene into the individual or
embryo to take over the function and compensate for the non-functional gene.
(c) First gene therapy was given to a four year old girl with Adenosine Deaminase (ADA)
deficiency by M Blease and WF Andresco in 1990s.
• ADA is caused due to the deletion of the gene for adenosine deaminase.
• In some children, ADA deficiency can be cured by bone marrow transplantation and enzyme
replacement therapy, but they are not completely curable.
(d) Steps involved are as follows:
• In first step of gene therapy, lymphocytes from the blood of the patient are grown in a culture
outside the body.
• A functional ADA cDNA (using a retroviral vector) is then introduced into these lymphocytes,
which are subsequently returned to the patient.
• As these cells are not immortal, the patient requires periodic infusion of such genetically
engineered lymphocytes.
• If the gene isolated from bone marrow cells producing ADA is introduced into cells at early
embryonic stages, it could be a permanent cure.
• Some other’diseases that can be treated by gene therapy are haemophilia, cystic fibrosis,
Parkinson’s disease, etc.
IV. Molecular diagnosis helps to solve the problem of early diagnosis and treatment of diseases.
(a) Using conventional methods of diagnosis (serum and urine analysis), early detection of
diseases is not possible.
(b) To overcome this problem, some molecular diagnosis techniques were developed that
provide early detection of diseases. These are as follows:
• Polymerase Chain Reaction (PCR) helps in early detection of diseases or pathogens by the
amplification of their nucleic acid.
Low concentration of pathogens (bacteria, viruses, etc) in the blood does not allow its detection.
PCR can amplify nucleic acids of such pathogens even when their concentration is very low.
PCR technique can be used for detecting HIV in suspected AIDS patients, genetic mutation in
suspected cancer patients and in identifying genetic disorders.
• Recombinant DNA technology is a modern molecular diagnostic technique. It is done in
the following steps:
A single stranded DNA or RNA tagged with a radioactive molecule called probe, is allowed to
hybridise to its complementary DNA in a clone of cells.
The cells are then detected by autoradiography.
The clone having mutated gene will not appear on the photographic film, because the probe will
not have complementarity with the mutated gene.
• Enzyme Linked Immuno Sorbent Assay (ELISA) is based on the principle of antigen-antibody
interaction. Infection by pathogen can be detected by the presence of antigens (proteins,
glycoproteins, etc) or by detecting the antibodies synthesised against the pathogen.