Very Short Answer Type Questions

Question. Why do children cured by enzyme-replacement therapy for adenosine deaminase deficiency need periodic treatment ? 
Answer : As this therapy does not cure the disease completely it requires periodic treatment. 
Detailed Answer :
They need periodic treatment because it is not a completely curative method. In this method, a functional ADA-cDNA is injected into the patient’s lymphocytes using a retroviral vector. Since, lymphocytes have a definite life cycle, there is a need for periodic infusion of genetically engineered lymphocytes (having ADA) into the patient. 

Question. State the role of C peptide in human insulin.
Answer : C-peptide is an extra stretch of polypeptide. It makes the insulin inactive. 

Question. What happens when Meloidogyne incognita consumes cells with RNAi gene ?
Answer : Using Agrobacterium vectors, the nematode specific genes are introduced into the host plant. The introduction of DNA initiates RNAi and thus silences the specific mRNA of Meloidogyne incognita.
As a consequence, the parasite can not survive in the transgenic host expressing specific interfering RNA. 

Question. Name the specific type of gene that is incorporated in a cotton plant to protect the plant against cotton boll worm infestation. 
Answer : Cry I Ac / Cry II Ab 

Question. List the type of cry genes that provide resistance to corn plants and cotton plants respectively against lepidopterans. 
Answer : cryIAc / cryIIAb – cotton. 
cryIAb – corn. 

Question. Name the technique by which Gene expression can be controlled with the help of RNA molecule.
OR
State a method of cellular defense which works in all eukaryotic organisms. 
Answer : RNA interference.

Question. Why do toxic insecticides proteins secreted by Bacillus thuringiensis kill insects ? 
Answer : It is because of Cry proteins produced by the spores of Bacillus thuringiensis which are toxic when ingested by some insects. 

Short Answer Type Questions

Question. Write the function of –
(i) Cry 1 AC gene
(ii) RNA interference (RNA)
Answer : (i) It produces inactive pro-toxin in the host cell / produces proteins to control cotton bollworms.
(ii) It produces dsRNA which silences host m-RNA / cellular defence mechanism / prevents infestation by nematodes. 

Question. Explain how Eli Lily an American company produced insulin by recombinant DNA technology. 
Answer : In 1983, Eli Lily and American company prepared two DNA sequences corresponding to A and B chains of human insulin, 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.

Question. Why is proinsulin called so ? How is insulin different from it ?
Answer : It is like a proenzyme or prohormone. It contains an extra stretch of C-peptide. It needs to be processed to become fully mature and functional hormone like insulin. 

Question. (i) State the role of DNA ligase in biotechnology.
(ii) What happens when Meloidogyne incognita consumes cells with RNAi gene ?
Answer : (i) Linking of DNA fragment is done by DNA ligase / linking of Okazaki fragments or discontinuous synthesis fragments / linking of desired gene with plasmid to form recombinant DNA.
(ii) Specific mRNA of the nematode silenced, parasite dies. 
Detailed Answer :
(i) The linking of antibiotic resistance gene with the plasmid vector became possible with the enzyme DNA ligase, which acts on cut DNA molecules and joins their ends. This makes a new combination of circular autonomously replicating DNA created in vitro and is known as recombinant DNA.
(ii) When Meloidogyne incognita (parasite) consumes cells with RNAi gene, parasite cannot survive and this prevents infestation. The introduced RNAi gene DNA forms both sense and anti-sense RNA.
Two strands being complementary to each other bind and form ds RNA, leading to RNAi. Thus, the mRNA of nematode is silenced and the parasite cannot survive there. 

Question. Why does a patient of ADA-deficiency requires repeated infusion of genetically engineered lymphocytes ? Suggest a possible permanent remedy. 
OR
Why is the introduction of genetically engineering lymphocytes into a ADA deficiency patient not a permanent cure ? Suggest a possible permanent cure. 
OR
Why is the introduction of genetically engineered lymphocytes into an ADA deficiency patient not a permanent cure ? Suggest a possible permanent cure. 
Answer : (i) ADA patients lacks functional T-lymphocytes and so fails to fight infectious pathogens.
(ii) The therapy includes reactivation of patient’s immune system by introduction of functional ADA gene.
(iii) The cells are not immortal and requires repeated infusions.
(iv) Introduction of ADA gene into early embryonic stages is the permanent cure.
Detailed Answer :
Adenosine deaminase (ADA) deficiency is associated with Severe Combined Immuno Deficiency (SCID).
(i) ADA is very crucial for the immune system to function. The deficiency of ADA causes severe combined immuno deficiency disease. The patient lack functional T-lymphocytes and fails to fight the infectious pathogens.
(ii) Using gene therapy, lymphocytes are extracted from the patient’s bone marrow and a normal functional gene for ADA is introduced into these lymphocytes with the help of the retrovirus.
(iii) The lymphocytes of bone marrow contain the functional ADA gene and reactivate the patient’s immune system.
(iv) 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 is that they are not completely curable. If the gene isolated from marrow cells producing ADA is introduced into cells at early embryonic stages, it could be a permanent cure.

Question. Explain how a hereditary disease can be corrected.
Give an example of first successful attempt made towards correction of such diseases.
Answer : Introduction of required genes into cells and tissues to treat diseases / by delivery of normal gene to take over the function of non-functional gene / by gene therapy. First gene therapy was given to four year old girl with Adenosine deaminase deficiency. 
Detailed Answer :
Gene therapy is an attempt to deal with hereditary or genetic or congenital diseases. This aims at correction of a genetic defect by delivery of a normal gene into an individual or embryo to take over or compensate the function for a non-functional gene. The first disease to have a gene therapy is ADA (Adenosine deaminase) deficiency. In this, the gene coding for enzyme ADA gets deleted leading to deficiency of ADA and problems in immune system. Gene therapy for ADA deficiency includes- Isolation of lymphocytes from patient’s blood and culturing them in-vitro.
Functional ADA cDNA are then introduced into the cultured lymphocytes.
These lymphocytes are returned back to the patient’s body. 
Permanent cure for this problem is the introduction of gene isolated from bone marrow cells producing ADA into cells at early embryonic stages. 

Question. How is Bt cotton made to attain resistance against bollworm ? 
Answer : (i) Specific Bt toxin genes were isolated from Bacillus thuringiensis and incorporated into the several crop plants such as cotton. The choice of genes depends upon the crop and the targeted pest, as most Bt toxins are insect-group specific.
(ii) The toxin is coded by a gene named cry. There are a number of them for example, the proteins encoded by the genes cryIAc and cryIIAb control the cotton ballworms and that of cryIAb controls corn borer. 

Question. Name the source and types of cry genes isolated from it for incorporation into crops by biotechnologies. Explain how have these genes brought beneficial change in the genetically modified crops ? 
Answer : Bacillus thuringiensis is the source of cry gene.
Types of crygenes isolated from it are cryIAc, cryIIAb, cryIAb.
These genes act as biopesticides when introduced.
They produce toxic insecticidal protein which, when activated cause death of the insects.

Question. Explain enzyme-replacement therapy to treat adenosine deaminase deficiency. Mention two disadvantages of this procedure.
Answer : Functional adenosine deaminase is given to the patient by injection. 
Disadvantages :
Therapy is not completely curative, periodic infusion of enzyme required. 
Detailed Answer :
Adenosine deaminase deficiency is a genetic disorder. The disorder is caused due to the deletion of the gene for adenosine deaminase, the enzyme crucial for the immune system to function.
Adenosine deaminase deficiency in patients can be treated by enzyme replacement therapy. In this treatment, patients are regularly injected with the functional ADA enzyme.
Disadvantages of this process :
(i) It does not completely eradicate the disease.
(ii) The requirement of repeated doses of the enzymes makes it expensive.

Question. Explain the various steps involved in the production of artificial insulin.
Answer : Two DNA sequences corresponding to A and B polypeptide chains of human insulin were prepared, these were introduced into E.coli to produce A and B chains separately, these chains were extracted and combined by creating disulphide bonds. 

Question. Name the organism from which the ‘cry‘ genes are isolated. Mention with the help of suitable example why and how bio-technologists have made use of ‘cry‘ genes ?
Answer : Bacillus thuringiensis 
(i) Source of insecticidal (crystal) protein that control the cotton bollworms / corn borer. 
(ii) Specific Bt toxin genes were isolated from Bacillus thuringiensis, incorporated into several crop plants such as cotton. 

Question. GM plants are useful in many ways. How would you convince farmers to grow GM plants on their field ? Explain giving three reasons.
Answer : Make crop more tolerant to abiotic stresses / Reduce reliance on chemical pesticides / Help to reduce post harvest loses / Increase efficiency of mineral usage / Enhance nutritional value of food.

Question. (i) What is Gene therapy?
(ii) Describe the procedure of such a therapy that could be a permanent cure for a disease. Name the disease. 
Answer : (i) Collection of methods that allows correction of gene defect that has been diagnosed in a child / embryo. Here, the genes are inserted into a person’s cells and tissues to treat a hereditary disease. It compensate the nonfunctional gene. This involves delivery of a normal gene into the individual / embryo to take over the function of non-functional / defective gene. 
(ii) If the desired gene is isolated and introduced into cells at early embryonic stages, it can provide a permanent cure. 
ADA/Adenosine deaminase deficiency. 

Question. A person is born with a hereditary disease with a weakened immune system due to deficiency of an enzyme. Suggest a technique for complete cure for this disease. Identify the deficient enzyme and explain the technique used for cure.
Answer : Gene Therapy.
ADA (Adenosine deaminase) deficiency.
Lymphocytes from the blood of the patient are grown in a culture, a functional ADA cDNA is introduced into these lymphocytes, which are subsequently returned to the patient. The permanent cure is done by introducing ADA cDNA into cells at early embryonic stages. 

Question. Explain how Eli Lily, an American company produced insulin by recombinant DNA technology.
OR
How did an American Company, Eli Lily use the knowledge of r-DNA technology to produce human insulin ?
OR
Explain how the company Eli lily was able to produce human insulin using rDNA technique.
OR
Recombinant DNA-technology is of great importance in the field of medicine. With the help of a flow chart, show how this technology has been used in preparing genetically engineered human insulins.
Answer : Prepared two DNA sequences, corresponding to A and B chains of human insulin, introduced in the plasmids of E.coli, to produce insulin chains, chains A and B were produced separately and extracted and combined by creating disulphide bonds 
Detailed Answer :
Preparation of human insulin using recombinant DNA technology :

Question. Rearrange the following in the correct sequence to accomplish an important biotechnological reaction : 
(a) In vitro synthesis of copies of DNA of interest
(b) Chemically synthesized oligonucleotides
(c) Enzyme DNA-polymerase
(d) Complementary region of DNA
(e) Genomic DNA template
(f) Nucleotides provided
(g) Primers
(h) Thermostable DNA-polymerase (from Thermus aquaticus)
(i) Denaturation of ds-DNA
Answer : 

Detailed Answer :
The correct sequence of the steps involved in biotechnological reaction is:
(e) Genomic DNA template
(i) Denaturation of ds-DNA
(g) Primers
(b) Chemically synthesised oligonucleotides
(c) Enzyme DNA polymerase
(h) Thermostable DNA polymerase
(f) Nucleotide provided
(d) Complementary region of DNA (a) In vitro synthesis of copies of DNA of interest 

Question. Name the nematode that damages the roots of tobacco plants. How is a transgenic tobacco plant made resistant to nematode using biotechnology ?
Answer : (i) Nematode – Meloidogyne incognita.
A nematode Meloidogyne incognita infect the roots of tobacco plants and causes a great reduction in yield. RNAi takes place in all eukaryotic organisms as a method of cellular defence. This method involves silencing of a specific mRNA due to a complementary dsRNA molecule that builds to and prevents translation of the mRNA (silencing).
(ii) Using Agrobacterium vectors, nematode-specific genes were introduced into the host plant. The introduction of DNA was such that it produces both sense and anti-sense RNA in the host cells.
These two RNAs being complementary to each other formed a double-stranded RNA (dsRNA) that initiated RNAi and thus silenced the specific mRNA of the nematode. As a consequence the parasite could not survive in a transgenic host expressing specific interfering RNA.

Question. (i) How has biotechnology helped in producing Meloidogyne incognita resistant tobacco plant ?
(ii) Why does this nematode die on eating such a GM plant ? 
Answer : (i) Nematodes like Meloidgyne incognitia infects the roots of tobacco plants and causes reduction in yield. The infestation of these nematodes can be prevented by the process of RNA interference (RNAi). RNAi is present in all eukaryotic organisms as cellular defence by silencing of specific mRNA due to complementary dsRNA molecules that bind to and prevents translation of the mRNA.
The source of complementary dsRNA may be from an infection by viruses having RNA genomes or mobile genetic elements that replicate through RNA intermediate.
(ii) Nematode specific genes were introduced into host plant using Agrobacterium vectors. The parasite could not survive in a transgenic host expressing specific interfering RNA.

Question. Expand ‘ELISA‘. Why is this method preferred over conventional method of diagnosis of disease ?
Answer : Enzyme Linked Immunosorbent Assay. 
Infection by pathogen detected by the presence of antigens (protein, glycoprotein etc.), antibodies synthesised against the pathogen. 
Conventional methods cannot provide early diagnosis which is made possible by ELISA. 

Question. Plasmid is a boon to biotechnology. Justify this statement quoting the production of human insulin as an example. 
Answer : The plasmids is a boon to biotechnology. It is a good vector in production of human insulin. It has a specific restriction site, where restriction endonuclease enzymes make a cut and a segment of DNA which codes for human insulin is inserted there. The recombinant plasmid so formed is introduced into E. coli and host cell where it replicates and produces insulin in large amount.
The plasmid has number of origin of replication (ori) where replication starts. 

Long Answer Type Questions

Question. Explain the application of rDNA technology to produce insulin.
Answer : Human insulin is synthesised as a pro-hormone.
The pro-hormone contains an extra C-peptide.
The C-peptide is not present in mature insulin and is removed during maturation. Eli-Lilyan 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. Chain A and B were produced separately, extracted and combined by creating disulphide bonds.

Question. (i) What is a plasmid ?
(ii) What is meant by ADA deficiency ? How is gene therapy a solution to this problem ? Why is it not a permanent cure ?
Answer : (i) Plasmids are extra-chromosomal, selfreplicating, usually circular double-stranded DNA molecules found in bacteria and in some yeast.
(ii) ADA is adenosine deaminase deficiency, this enzyme is crucial for the immune system to function. The patient lacks functional T-lymphocytes and fails to fight the infecting pathogens.
Children with ADA deficiency are cured by bone marrow transplantation or enzyme replacement therapy, where ADA is given by injection. By using gene therapy techniques, lymphocytes are taken from the patient’s bone marrow and the normal gene for ADA is introduced into the lymphocytes using retrovirus. These cells are re-introduced in the patient’s immune system.
As these cells are not immortal, the patient requires periodic infusion of such genetically engineered lymphocytes. Hence, it is not a permanent cure.
If the functional gene is introduced into the bone marrow cells at early embryonic stage, it would be a permanent cure.