12.

BIOTECHNOLOGY AND ITS

APPLICATIONS

Multiple Choice Questions

Single Correct Answer Type
1. Bt cotton is not
(a) A GM plant
(b) Insect resistant
(c) A bacterial gene expressing system
(d) Resistant to all pesticides
Answer. (d) Bt cotton is a GM plant, insect resistant and a bacterial gene expressing system.

2. C-peptide of human insulin is

(a) A part of mature insulin molecule
(b) Responsible for formation of disulphide bridges
(c) Removed during maturation of pro-insulin to insulin
(d) Responsible for its biological activity.
Answer. (c) C-peptide of human insulin is removed during maturation of pro-insulin to insulin.

3. GEAC stands for

(a) Genome Engineering Action Committee
(b) Ground Environment Action Committee
(c) Genetic Engineering Approval Committee .
(d) Genetic and Environment Approval Committee

Answer. (c) GEAC stands for Genetic Engineering Approval Committee.

4. α -1 antitrypsin is
(a) An antacid
(b) An enzyme
(c) Used to treat arthritis
(d) Used to treat emphysema.
Answer. (d) α -1 antitrypsin is used to treat emphysema.

5. A probe which is a molecule used to locate specific sequences in a mixture of, DNA or
RNA molecules could be
(a) A single stranded RNA
(b) A single stranded DNA
(c) Either RNA or DNA
(d) Can be ssDNA but not ssRNA
Answer. (c) A single stranded DNA or RNA, tagged with a radioactive molecule is called Probe.
.
6. Choose the correct option regarding Retrovirus:
(a) An RNA virus that can synthesise DNA during infection
(b) A DNA virus that can synthesise RNA during infection
(c) A ssDNA virus
(d) AdsRNAvirus
Answer. (a) Retrovirus is an RNA virus that can synthesise DNA during infection.

7. The site of production of ADA in the body is

(a) Erythrocytes ‘
(b) Lymphocytes
(c) Blood plasma
(d) Osteocytes
Answer. (b) The site of production of ADA in the body is lymphocytes.

8. A protoxin is
(a) A primitive toxin .
(b) A denatured toxin
(c) Toxin produced by protozoa
(d) Inactive toxin
Answer. (d) Bt toxin is a protein. This toxin does not kill the Bacillus because 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.

9. Pathophysiology is the
(a) Study of physiology of pathogen
(b) Study of normal physiology of host
(c) Study of altered physiology of host
(d) None of the above
Answer. (c) Pathophysiology is the study of altered physiology of host.

10. The trigger for activation of toxin of Bacillus thuringiensis is

(a) Acidic pH of stomach

(b) High temperature

(c) Alkaline pH of gut
(d) Mechanical action in the insect gut
Answer. (c) The trigger for activation of-toxin of Bacillus thuringiensis is alkaline pH of gut.
11. Golden rice is
(a) A variety of rice grown along the yellow river in China
(b) Long stored rice having yellow colour tint
(c) A transgenic rice having gene for b-carotene
(d) Wild variety of rice with yellow coloured grains.
Answer. (c) Golden rice is a transgenic rice having gene for β-carotene.

12. In RNAi, genes are silenced using

(a) ss DNA (b) ds DNA
(c) dsRNA (d) ssRNA
Answer. (c) RNAi takes place in all eukaryotic organisms as a method of cellular defense. This
method involves silencing of a specific mRNA due to the complementary dsRNA molecule
that binds to and prevents translation of the mRNA (silencing).

13. The first clinical gene therapy was done for the treatment of
(a) AIDS . (b) Cancer
(c) Cystic fibrosis
(d) SCID (Severe Combined Immuno Deficiency resulting from deficiency of ADA)
Answer. (d) A first clinical gene therapy was given in 1990 to a 4 year old girl with ADA
(Adenosine deaminase) deficiency. ADA deficiency causes SCID (Severe Combined Immuno
Deficiency) in which B-lymphocytes and T-lymphocytes are not formed. .

14. ADA is an enzyme which i§ deficient in a genetic disorder SCID. What is the full form of
ADA?
(a) Adenosine deoxy aminase (b) Adenosine deaminase
(c) Aspartate deaminase (d) Arginine deaminase
Answer. (b) ADA stands for Adenosine deaminase.

15. Silencing of a gene could be achieved through the use of

(a) (RNAi) only .(b) Antisense RNA only
(c) By both (d) None of the above
Answer. (c) Silencing of a gene could be achieved through the use of (RNAi) only and
Antisense RNA only.

Very Short Answer Type Questions

1. In view of the current food crisis, it is said, that we need another green revolution.
Highlight the major limitations of the earlier green revolution.
Answer. Limitations of green revolution:

1. 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 (fertilisers 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.
2. Use of agrochemicals causes eutrophication in aquatic ecosystems and
biomagnification in terrestrial ecosystems.
3. Water logging and soil salinity are some of the problems that have come in the wake of
green revolution.
2. Expand GMO. How is it different from a hybrid?
Answer. GMO stands for Genetically Modified Organism. It differs from a hybrid because in a
hybrid,cross is done between total genomes of two species or strains, where as in a GMO,
foreign gene(s) is introduced in the organism and is usually maintained as extra-
chromosomal entity or is integrated into the genome of the organism. .

3. Differentiate between diagnostics and therapeutics. Give one example and for each
category.
Answer. A diagnostic technique helps us to identify a disease. Example: ELISA is a test for
HIV.
A therapeutic agent on the other hand helps in the treatment of a disease. Example:
Antibiotics for bacterial infections.

4. Give the full form of ELISA. Which disease can be detected using it? Discuss the principle
underlying the test.
Answer. Enzyme Linked Immuno-sorbent Assay (ELISA) is used for the detection of AIDS.
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.

5. Can a disease be detected before its symptoms appear? Explain the principle involved.
Answer. Presence of a pathogen (bacteria, viruses, etc.) is normally suspected only when the
pathogen has produced a disease symptom. By this time the concentration of pathogen is
already very high in the body. However, very low concentration of a bacteria or virus (at a time
when the symptoms of the disease are not yet visible) can be detected by amplification of
their nucleic acid by PCR.

6. Write a short note on biopiracy highlighting the exploitation of developing countries by the
developed countries.
Answer. Biopiracy is the term used to refer to the use of bio-resources by multinational
companies and other organisations without proper authorisation from the countries and
people concerned without compensatory payment. Most of the industrialised nations are rich
financially but poor in biodiversity arid traditional knowledge. In contrast the developing and
the underdeveloped world is rich in biodiversity and traditional knowledge related to bio-
resources. Traditional knowledge related to bio-resources can be exploited to develop modem
applications and can also be used to save time, effort and expenditure during their
commercialisation.

7. Many proteins are secreted in their inactive form. This is also hue of many toxic proteins
produced by micro organisms. Explain how the mechanism is useful for the organism
producing the toxin.
Answer. Many proteins including certain toxins are secreted in their inactive form. They get
activated, only when exposed to a specific trigger (pH. temperature etc.). It is advantageous to
the bacteria producing it because the bacteria does not get killed due to the action of protein.

8. While creating genetically modified organisms, genetic barriers are not respected. How
can this be dangerous in the long run?
Answer. The manipulation of living organisms by the human race cannot go on any further,
without regulation. Some ethical standards are required to evaluate the morality of all human
activities that might help or harm living organisms. Going beyond the morality of such issues,
the biological significance of such things is also important. Genetic modification of organisms
can have unpredictable results when such organisms are introduced into the ecosystem.

9. Why has the Indian Parliament cleared the second amendment of the country’s patents
bill?
Answer. The Indian Parliament has recently cleared the second amendment of the Indian
Patents Bill, that take such issues into consideration, including patent terms emergency
provisions and research and development initiative.

10. Give any two reasons why the patent on Basmati should not have gone to an American
Company.
Answer. (i) Rice is an important food grain, the presence of which goes back ‘ thousands of
years in Asia’s agricultural history. There is an estimated 200,000 varieties of rice in India
alone. The diversity of rice in India is one of the richest in the world. Basmati rice is distinct for
its unique aroma and flavour and 27 documented varieties of Basmati are grown in India.
(ii) There is a reference to Basmati in ancient texts, folklore and poetry.

11. How was Insulin obtained before the advent of rDNA technology? What were the
problems encountered?
Answer. Insulin used for diabetes was earlier extracted from pancreas of slaughtered cattle
and pigs. Insulin from an animal source, though caused some patients to develop allergy or
other types of reactions to the foreign protein.

12. With respect to understanding diseases,’discuss the importance of transgenic animal

models.
Answer. Many transgenic animals are designed to increase our understanding of how genes
contribute to 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.

13. Name the first transgenic cow. Which gene was introduced in this cow?
Answer. Rosie was the name of the first transgenic cow. Gene for human alpha lactalbumin
was introduced in its gene, which made the milk nutritionally richer.

14. PCR is a useful tool for early diagnosis of an infectious disease. Elaborate.
Answer. PCR is a very sensitive technique which enables the specific amplification of desired
DNA from a limited amount of DNA template. Hence, it can detect the presence of an
infectious organism in the infected patient at an early stage of infection (even before the
infectious organism has multiplied to large number).

15. What is GEAC and what are its objectives?

Answer. GEAC (Genetic Energy Approval Committee) is an Indian government organisation.
Its objective are to:
(a) examine the validity of GM (Genetic modification of organism) research.
(b) inspect the safety of introducing GM for public services.

16. For which variety of Indian rice, the patent was filed by a USA Company?
Answer. Indian Basmati was crossed with semi-dwarf variety and was claimed as a new
variety for which the patent was filed by a USA company.

17. Discuss the advantages of GMO.

Answer. 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:

1. Made crops more tolerant to abiotic stresses (cold, drought, salt, heat).
2. Reduced reliance on chemical pesticides (pest-resistant crops).
 3. Helped to reduce post-harvest losses.
4. Increased efficiency of mineral usage by plants (this prevents early exhaustion of fertility
of soil).
5. Enhanced nutritional value of food, e.g., Vitamin ‘A’ enriched rice.

Short Answer Type Questions

1. Gene expression can be controlled with the help of RNA. Explain the method with an
example.
Answer. Gene expression can be controlled by using RNA molecule. The technology is called
RNA interference or RNAi. It is used to block the expression of certain genes and also referred
to as gene silencing. During this process a complementary RNA to the mRN A being produced
by the gene is introduced into the cell. This RNA binds to the mRNA making it double stranded
and therefore stops translation. Resistance to nematode Meloidegyne incognita in tomato has
been achieved by this method.

2. Ignoring our traditional knowledge can we prove costly in the area of biological patenting.
Justify.
Answer. Most of the industrialised nations are rich financially but poor in biodiversity and
traditional knowledge. In contrast the developing and the underdeveloped world is rich in
biodiversity and traditional knowledge related to bio-resources. Traditional knowledge related
to bio-resources can be exploited to develop modem applications and can also be used to
save time, effort and expenditure during their commercialisation. There has been” growing
realisation of the injustice, inadequate compensation and benefit sharing between developed
and developing countries. Therefore, some nations are developing laws to prevent such
unauthorised exploitation of their bio-resources and traditional knowledge.

3. Highlight any four areas where genetic modification of plants has been useful.
Answer. 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
(i) Made crops more tolerant to abiotic stresses (cold, drought, salt, heat).
(ii) Reduced reliance on chemical pesticides (pest-resistant crops).
(iii) Helped to reduce post-harvest losses.
(iv) Increased efficiency of mineral usage by plants (this prevents early exhaustion of fertility
of soil).

4. What is a recombinant DNA vaccine? Give two examples.

Answer. A recombinant vaccine is a vaccine produced through recombinant DNA technology.
This involves inserting the DNA encoding an antigen that stimulates an immune response into
bacterial or mammalian cells. Recombinant DNA technology has allowed the production of
antigenic polypeptides of pathogen in bacteria or yeast. Vaccines produced using this
.approach allow large scale production and hence greater availability for immunisation, e.g.,
hepatitis B vaccine (Recombivax HB) produced from yeast. As of June 2015 one human DNA
vaccine had been approved for human use, the single-dose Japanese encephalitis vaccine
called IMOJEV, released in 2010 in Australia.

5. Why is it that the line of treatment for a genetic disease is different from infectious
diseases?
Answer. If a person is born with a hereditary disease, can a corrective therapy be taken for
such a disease? Gene therapy is an attempt to do this. Gene therapy is a collection of
methods that allows correction of a gene defect that has been diagnosed in a child/embryo.

6. Discuss briefly how a probe is used in molecular diagnostics.

Answer. A single stranded DNA or RNA, tagged with a radioactive molecule (probe) is allowed
to hybridise to its complementary DNA in a clone of cells followed by detection using
autoradiography. The clone having the mutated gene will hence not appear on the
photographic film, because the probe will not have complementarity with the mutated gene.

7. Who was the first patient who was given gene therapy? Why was the given treatment
recurrent in nature?
Answer. The first clinical gene therapy was given in 1990 to a 4-year old girl with adenosine
deaminase (ADA) deficiency. This enzyme is crucial for the immune system to function. The
disorder is caused due to the deletion of the gene for adenosine deaminase. In some children
ADA deficiency can be cured by bone marrow transplantation; in others it can be treated by –
enzyme replacement therapy, in which functional ADA is given to the patient by injection. But
the problem- with both of these approaches that they are not completely curative. As a first
step towards 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. However, as these cells
are not immortal, the patient requires “periodic infusion of such genetically engineered
lymphocytes. However, if the gene isolate from marrow cells producing ADA is introduced into
cells at early embryonic stages, it could be a permanent cure.

8. Taking examples under each category, discuss upstream and downstream processing.
Answer. Upstream processing: Biotechnological processes can be separated into upstream
processes and dpwnstream processes. The upstream process is defined as the entire
process from DNA isolation and culture expansion of the cells until final product.
Downstream processing: After completion of the biosynthetic stage, the product has to be
subjected through a series of processes before it Is ready for marketing as a finished product.
The processes include separation and purification, which are collectively referred to as
downstream processing. The product has to be formulated with suitable preservatives. Such
formulation has to undergo through clinical trials as in case of drugs. Strict quality control
testing for each product is also required. The downstream processing and quality control
testing vary from product to product.

9. Define Antigen and Antibody. Name any two diagnostic kits based upon them.
Answer. An antigen is a foreign substance that elicits the formation of an antibody. Antibody
is a protein that is synthesised in response to an antigen. Antigen and antibody show high
degree of specificity in binding each other. Two diagnostic kits based on antigen-antibody
interaction are:
(a) ELISA for HIV.
(b) Pregnancy test kits.

10. ELISA technique is based on the principles of antigen-antibody interaction. Can this
techique be used in the molecular diagnosis of a genetic disorder, such as phenyketonuria?
Answer. Yes. One can use antibody against the enzyme (that is responsible for the
metabolism of phenylalanine) to develop ELISA based diagnostic technique. The patient
where the enzyme protein is absent would give negative result in ELISA when compared to
normal individual.

11. How is a mature, functional insulin hormone different from its prohormone form?
Answer. Mature functional insulin is obtained by processing of pro-hormone which contains
extra peptide called C-peptide. This C-peptide is removed during maturation of pro-insulin to
insulin.

12. Gene therapy is an attempt to correct a genetic defect by providing a normal gene into
the individual. By this’the normal function can be restored. An alternate method would be to
provide the gene product (protein/enzyme) known as enzyme replacement therapy, which
would also restore the function. Which in your opinion is a better option? Give reason for
your answer.
Answer. Gene therapy is an attempt to correct a genetic defect by providing a normal gene
into the individual. By this the normal function can be restored. Alternate method would be to
provide the gene product (protein/ enzyme) know as enzyme replacement therapy, which
would also restore the function. Which in your opinion is a better option? Give reason for your
answer.

13. Transgenic animals are the animals in which a foreign gene is expressed. Such animals
can be used to study the fundamental biological process, phenomenon as well as for
producing products useful for mankind. Give one example for each type.
Answer. Study of basic biological process—how genes are regulated, how they affect the
normal functions of the body and its development. Transgenic cow, Rosie is an example for
the second category.

14. When a foreign DNA is introduced into an organism, how is it maintained in the host and
how is it transferred to the progeny of the organism?
Answer. Foreign gene is usually ligated to a plasmid vector and introduced in the host. As
plasmid replicates, and makes multiple copies of itself, so does the foreign gene gets
replicated and its copes are made. When the host organism divides, its progeny also receives
the plasmid DNA containing the foreign gene.

15. Bt cotton is resistant to pest, such as lepidopteron, dipterans and coleopterans. Is Bt

cotton also resistant to other pests as well?
Answer. Bt cotton is made resistant to only certain specific taxa of pests. It is quite likely that
in future,some other pests may infest this Bt cotton plants. It has similar immunisation
against small-pox which does not provide immunity against other pathogens like those that
cause cholera, typhoid etc.

Long Answer Type Questions

1. A patient is suffering from ADA deficiency. Can he be cured. How?
Answer. The first clinical gene therapy was given in 1990 to a 4-year old girl with adenosine
deaminase (ADA) deficiency. This enzyme is crucial for the immune system to function. The
disorder is caused due to the deletion of the gene for adenosine deaminase. In some children
ADA deficiency can be cured by bone marrow transplantation; in others it can be treated by
enzyme replacement therapy, in which functional ADA is given to the patient by injection. But
the problem with both of these approaches that they are not completely curative. As a first
step towards 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. However, as these cells
are not immortal, the patient requires periodic infusion of such genetically engineered
lymphocytes. However, if the gene isolate from marrow cells producing ADA is introduced into
cells at early embryonic stages, it could be a permanent cure.

2. Define transgenic animals. Explain in detail any four areas where they can be utilised.
Answer. Transgenic animals are the products of genetic engineering and express specific
gene(s) from totally unrelated source. Following are the four main areas where they can be
utilised: .
(1) To study normal physiology and development these animals can be used to study as to
which factor/gene products are needed at what time of development. By expression of certain
genes, they help scientists to understand the normal gene expression at various stages of
growth and development. 4
(2) Study of Diseases: Transgenic animals can be created to serve as models for various
human diseases. They also help us understand the involvement of various genes in diseases
like cancer, Parkinson’s disease etc.
(3) Vaccine safety: Transgenic animals can be used to test vaccines like polio vaccine.
Transgenic mice have shown promising results in this area and would replace the vaccine
testing on monkeys in the years to come.
(4) Chemical safety testing: Transgenic animals are created which are more sensitive to
certain chemicals/drugs. These are used to study the toxicity or side effects of that
chemical/drug. The advantage is that we get results faster.

3. You have identified a useful gene in bacteria. Make a flow chart of the steps that you
would follow to transfer this gene to a plant.
Answer. After identifying a useful gene in bacteria, following steps should be undertaken:

4. Highlight five areas where biotechnology has influenced our lives.

Answer. The applications of biotechnology include:

1. therapeutics and diagnostics

2. genetically modified crops for agriculture
3. processed food
4. bioremediation
5. waste treatment and energy production.

5. What are the various advantages of using genetically modified plants to increase the
overall yield of the crop?
Answer. GM plants have been useful in many ways. Genetic modification has:

1. Made crops more tolerant to abiotic stresses (cold, drought, salt, heat).
2. Reduced reliance on chemical pesticides (pest-resistant crops).
3. Helped to reduce post-harvest losses.
4. Increased efficiency of mineral usage by plants (this prevents early exhaustion of fertility
of soil).
5. Enhanced nutritional value of food>e.g., Vitamin ‘A’ enriched rice.

In addition to these uses, GM has been used to create tailor-made plants to supply alternative
resources to industries, in the form of starches, fuels and pharmaceuticals.

6. Explain with the help of one example how genetically modified plants can:
(a) Reduce usage of chemical pesticides .
(b) Enhance nutritional value of food crops
Answer. (a) Reduce usage of chemical pesticides: Bt toxin is produced by a bacterium called
Bacillus thuringiensis (Btfor short). Bt toxin gene has been cloned from the bacteria and been
expressed in plants to provide resistance to insects without the need for insecticides; in effect
created a bio-pesticide. Examples are Bt cotton, Bt com, rice, tomato, potato and soyabean
etc.
(b) Enhance nutritional value of food crops: Golden rice is the transgenic variety of basmati
rice which gives high yield and rich in vitamin A,
so it is used in the deficiency of vitamin-A causing night blindness and skin disorder.

7. List the disadvantages of insulin obtained from the pancreas of slaughtered cows and
pigs:
Answer.

1. Insulin being a hormone is produced in very little amounts in the body.

Hence, a large number of animals need to be sacrificed for obtaining small quantities of
insulin. This makes the cost of insulin very high. [Demand being many fold higher than
supply].
2. Slaughtering of animals is also not ethical.
3. There is potential of immune response in humans against the administered insulin
which is derived from animals.
4. There is possibility of slaughtered animals being infested with some infectious micro
organism which may contaminate insulin.

8. List the advantages of recombinant insulin.

Answer. Insulin used for diabetes was earlier extracted from pancreas of slaughtered cattle
and pigs. Insulin from an animal source, though caused some patients to develop allergy or
other types of reactions to the foreign protein. Insulin consists of two short polypeptide
chains: chain A and chain B, that are linked together by disulphide bridges. In mammals,
including humans, insulin is synthesised as a pro-hormone (like a pro-enzyme, the pro-
hormone also needs to be processed before it becomes a fully mature and functional
hormone) which contains an extra stretch called the C peptide. This C peptide is not present
in the mature insulin and is removed during maturation into insulin. The main challenge for
production of insulin using rDNA techniques was getting insulin assembled into a mature
form. In 1983, Eli Lilly an American company 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
disulfide bonds to form human insulin.

9. What is meant by the term bio-pesticide? Name and explain the mode of action of a
popular bio-pesticide.
Answer. Biopesticide is a pesticide which is
(a) not chemical in nature
(b) more specific in action against the pest
(c) safer for environment than chemical pesticides
A popularly known bio-pesticide is Bt toxin, which is produced by a bacterium called Bacillus
thuringiensis. Bt toxin gene has been cloned from this bacterium and expressed in plants. Bt
toxin protein when ingested by the insect, gets converted to its active form due to the alkaline
pH of the gut. The activated toxin binds to the surface of midgut epithelial cells and create
pores that cause cell swelling and lysis and eventually kills the insect.

10. Name the five key tools for accomplishing the tasks of recombinant DNA technology.
Also mention the functions of each tool.
Answer.

1. Restriction endonucleases: for cutting the desired DNA at desired places

2. Gel electrophoresis: for separating the desired DNA fragments
3. Ligase enzyme: for creating recombinant DNA molecule.
4. DNA delivery system: like electroporation, microinjection, gene gun method.
5. Competent host (usually bacteria/yeast): to take up recombinant DNA.