Biotechnology and Its Applications: Class 12 Biology Practice Questions

Dr. Priya, a molecular biologist at a research institute in Pune, is working on producing a therapeutic protein — human insulin — using recombinant DNA technology. She isolates the human insulin gene from a cDNA library and clones it into plasmid pMI322 (which carries amp^R and tet^R genes) by inserting the insulin gene within the tet^R gene using EcoRI. Transformed E. coli cells are plated on ampicillin-only and tetracycline-only plates. Plate A (ampicillin) shows many colonies; Plate B (tetracycline) shows very few colonies.

Read the following passage carefully and answer the questions that follow:

Dr. Priya, a molecular biologist at a research institute in Pune, is working on producing a therapeutic protein — human insulin — using recombinant DNA technology. She isolates the human insulin gene (comprising A-chain and B-chain coding sequences) from a cDNA library. She clones this gene into a plasmid vector called pMI322, which contains an ampicillin resistance gene (amp^R) and a tetracycline resistance gene (tet^R). The insulin gene is inserted within the tet^R gene using the restriction enzyme EcoRI. The recombinant plasmid is then introduced into E. coli cells by treating them with CaCl₂ solution followed by heat shock. The transformed bacteria are plated on two sets of agar plates — one containing ampicillin only, and another containing tetracycline only.

Dr. Priya observes the following results after overnight incubation:

| Plate | Antibiotic | Colonies observed |
|-------|------------|------------------|
| Plate A | Ampicillin only | Many colonies |
| Plate B | Tetracycline only | Very few colonies |

She then transfers the bacteria to a large stirred-tank bioreactor for large-scale production of insulin.

(a) Why did Dr. Priya use cDNA rather than genomic DNA to clone the human insulin gene into E. coli? (1 mark)

(b) Identify which colonies on Plate A contain the recombinant plasmid (with the insulin gene inserted) and which contain the non-recombinant plasmid. Justify your answer using the principle of insertional inactivation. (2 marks)

(c) Name ONE essential feature that the plasmid pMI322 must possess (other than antibiotic resistance genes) to ensure successful cloning, and state its function. (1 mark)

A biotechnology company was developing a genetically modified variety of brinjal (Bt brinjal) to protect it from the fruit and shoot borer — a major pest that causes significant crop loss. Scientists introduced a specific gene from the soil bacterium Bacillus thuringiensis into the brinjal genome. When the insect larvae feed on the modified plant, the protein encoded by the introduced gene is activated in the alkaline pH of the insect's midgut, creating pores in the epithelial cells, leading to the death of the larva. Before releasing this GM crop commercially in India, the company had to obtain approval from a statutory body.

Read the following passage and answer the questions that follow:

A biotechnology company was developing a genetically modified variety of brinjal (Bt brinjal) to protect it from the fruit and shoot borer — a major pest that causes significant crop loss. Scientists introduced a specific gene from the soil bacterium Bacillus thuringiensis into the brinjal genome. When the insect larvae feed on the modified plant, the protein encoded by the introduced gene is activated in the alkaline pH of the insect's midgut, creating pores in the epithelial cells, leading to the death of the larva. Before releasing this GM crop commercially in India, the company had to obtain approval from a statutory body.

(i) Name the gene introduced into brinjal from Bacillus thuringiensis that confers resistance against the fruit and shoot borer. (1 mark)

(ii) Explain the mechanism by which the protein encoded by this gene kills the insect larva. (2 marks)

(iii) Name the statutory body in India that is responsible for approving the release of genetically modified organisms for commercial use. (1 mark)

A biotechnology company is producing human insulin using recombinant DNA technology. Scientists isolated the insulin gene from human pancreatic cells and inserted it into a plasmid vector. The recombinant plasmid was then introduced into Escherichia coli bacteria. The bacteria were grown in large fermenters, and the insulin protein was extracted and purified for medical use. Before the availability of this recombinant human insulin, diabetic patients were treated with insulin extracted from the pancreas of slaughtered cattle and pigs, which sometimes caused allergic reactions in patients.

Read the following passage and answer the questions that follow:

A biotechnology company is producing human insulin using recombinant DNA technology. Scientists isolated the insulin gene from human pancreatic cells and inserted it into a plasmid vector. The recombinant plasmid was then introduced into Escherichia coli bacteria. The bacteria were grown in large fermenters, and the insulin protein was extracted and purified for medical use. Before the availability of this recombinant human insulin, diabetic patients were treated with insulin extracted from the pancreas of slaughtered cattle and pigs, which sometimes caused allergic reactions in patients.

(a) Name the type of plasmid used as a vector in recombinant DNA technology and state ONE property that makes it suitable as a vector. (1 mark)

(b) Draw a labelled diagram showing the structure of a recombinant plasmid after insertion of the human insulin gene. Label any THREE essential features. (2 marks)

(c) State ONE advantage of using recombinant human insulin over animal-derived insulin for treating diabetes. (1 mark)

A biotechnology company is developing a genetically engineered bacterium to produce human insulin. The team extracts mRNA from human pancreatic beta cells and uses reverse transcriptase to synthesise cDNA. This cDNA is then inserted into a plasmid vector (pBR322) and introduced into Escherichia coli. However, the team faces two challenges:

Challenge 1: When the foreign gene is inserted into the tetracycline-resistance gene (tet^R) of pBR322, the scientists need a reliable method to identify which bacterial colonies have successfully taken up the recombinant plasmid.

Challenge 2: The team realises that if they had used the original genomic DNA (with introns) from human cells instead of cDNA, the bacteria would NOT have produced functional insulin.

Read the passage given below and answer the questions that follow:

A biotechnology company is developing a genetically engineered bacterium to produce human insulin. The team extracts mRNA from human pancreatic beta cells and uses reverse transcriptase to synthesise cDNA. This cDNA is then inserted into a plasmid vector (pBR322) and introduced into Escherichia coli. However, the team faces two challenges:

Challenge 1: When the foreign gene is inserted into the tetracycline-resistance gene (tet^R) of pBR322, the scientists need a reliable method to identify which bacterial colonies have successfully taken up the recombinant plasmid.

Challenge 2: The team realises that if they had used the original genomic DNA (with introns) from human cells instead of cDNA, the bacteria would NOT have produced functional insulin.

(a) Why did the scientists use mRNA from pancreatic beta cells to make cDNA, rather than using genomic DNA directly? Give ONE reason. [1]
(b) Explain the principle of insertional inactivation used in Challenge 1 to identify recombinant colonies. [2]
(c) Name the enzyme used to synthesise cDNA from mRNA, and state ONE property of this enzyme that makes it unique. [1]

A biotechnology company is working on producing human insulin for diabetic patients. Scientists isolated the human insulin gene and inserted it into a plasmid vector. The recombinant plasmid was then introduced into Escherichia coli bacteria. The bacteria were grown in large fermentation tanks (bioreactors) to produce insulin on an industrial scale. The insulin produced was then extracted, purified, and made available as 'Humulin' for patients.

Read the following passage and answer the questions that follow:

A biotechnology company is working on producing human insulin for diabetic patients. Scientists isolated the human insulin gene and inserted it into a plasmid vector. The recombinant plasmid was then introduced into Escherichia coli bacteria. The bacteria were grown in large fermentation tanks (bioreactors) to produce insulin on an industrial scale. The insulin produced was then extracted, purified, and made available as 'Humulin' for patients.

(a) Name the type of insulin produced by this method and state ONE advantage it has over insulin extracted from animals. (1 mark)
(b) Name the two polypeptide chains that make up the human insulin molecule. Why was human insulin produced as two separate chains in bacteria and then combined? (2 marks)
(c) Name any TWO features that the plasmid used as a vector in the above process must possess. (2 marks)

Assertion (A): In the production of recombinant insulin, the A and B chains are produced separately in E. coli and then combined to form functional insulin.
Reason (R): Eukaryotic genes expressed in bacteria produce mRNA that cannot be translated without introns being removed by the bacterial machinery.

A farmer in Punjab grows Bt cotton (carrying the cry1Ac gene) to protect against bollworm. However, he notices that the same crop is severely damaged by whitefly, a phloem-sucking insect. He concludes that Bt cotton cannot protect against all types of insect pests. Which of the following best explains why Bt cotton fails to control the whitefly?

(A) The whitefly does not feed on plant tissue, so it is never exposed to the Bt toxin.
(B) The whitefly's midgut is acidic, so the inactive protoxin is not converted to its active toxic form.
(C) The whitefly lacks the specific receptors on its midgut epithelial cells that bind the activated Bt toxin.
(D) The cry1Ac gene is expressed only in the roots of Bt cotton and not in the phloem.

A farmer notices that his cotton crop is being severely damaged by bollworm infestation every season. His neighbour, who planted Bt cotton, has no such problem. Which of the following correctly explains why Bt cotton is resistant to bollworm?

A researcher isolates a gene from a thermophilic bacterium and wants to express it in E. coli to produce the encoded enzyme commercially. She constructs a recombinant plasmid and transforms E. coli cells. However, the enzyme produced in E. coli is non-functional. Which of the following is the MOST likely reason for this outcome?

A farmer notices that his Bt cotton crop is resistant to bollworm attack, while the neighbouring non-Bt cotton crop is heavily damaged. The reason for this resistance in Bt cotton is that it produces a toxin derived from:

A crop scientist observes that a particular transgenic cotton variety suffers no damage from bollworm larvae, even though the adult bollworm moths can lay eggs on its leaves. She notes that the plant produces a protein that is inactive at neutral pH but becomes toxic only inside the alkaline gut of the bollworm larva. Which of the following conclusions is best supported by this observation?

A scientist inserts a foreign gene into the tetracycline-resistance gene (tet^R) of the plasmid pBR322 and transforms it into E. coli. The bacterial colonies are then replica-plated onto two plates: one containing ampicillin and one containing tetracycline. (i) On which plate will the recombinant colonies FAIL to grow? (ii) Give ONE reason why.

Draw a labelled diagram of the stirred-tank bioreactor used for large-scale production of biological products. Label any two important structural features.

Distinguish between 'Insertional inactivation' and 'Antibiotic resistance' as selectable markers used in recombinant DNA technology. Give one example of each.

Draw a well-labelled diagram of the lac operon in the ABSENCE of lactose (repressed state). Name the molecule that acts as the inducer of the lac operon and state what happens to the operon when this inducer is present.

A biotech company has developed a genetically modified (GM) brinjal variety by introducing a cry1Ab gene from Bacillus thuringiensis into its genome. Before this variety can be released for commercial cultivation in India, it must pass through a mandatory regulatory body.

(i) Name the regulatory body that approves the release of GM crops for commercial use in India.
(ii) The cry1Ab gene produces a protoxin inside the transgenic brinjal plant. Explain how this protoxin becomes lethal specifically to the target insect pest (fruit and shoot borer) but does NOT harm humans who eat the brinjal.
(iii) A farmer argues: 'If the Bt brinjal plant already produces the insecticidal protein in every cell, why should I still follow Integrated Pest Management (IPM) practices?' Give ONE scientifically valid reason to counter this argument.

A biotechnology company developed a transgenic tobacco plant by introducing a nematode-specific gene using Agrobacterium tumefaciens as a vector. The gene introduced produces double-stranded RNA (dsRNA) in the plant cells. When the nematode Meloidogyne incognita feeds on the roots of this plant, it dies, but the tobacco plant itself is unharmed.

(i) Name the molecular mechanism by which the nematode is killed after ingesting the dsRNA.
(ii) Identify the specific enzyme that is activated inside the nematode cell and describe its role in silencing the nematode's own gene.
(iii) Why does this mechanism NOT harm the tobacco plant itself, even though the same dsRNA is present in its cells?

Explain the steps involved in the production of a recombinant protein using a bioreactor. Name any one protein produced by this method and state its use.

A biotechnology company is developing a recombinant insulin product. The gene encoding human insulin (preproinsulin) was isolated from a human pancreatic cell. However, when this genomic DNA was directly inserted into an E. coli expression system, no functional insulin protein was produced.

(i) Explain why the direct use of genomic DNA failed to produce functional insulin in E. coli. (1 mark)

(ii) Suggest the correct form of DNA that should be used and describe how it is prepared. (1 mark)

(iii) Draw a labelled diagram showing the steps involved in the preparation of this correct form of DNA from mRNA. (1 mark)

A pharmaceutical company is producing human insulin using recombinant DNA technology. The gene for human insulin was isolated from human pancreatic cells and inserted into a bacterial expression system. However, the scientists found that when the same insulin gene was directly cloned from genomic DNA and expressed in E. coli, the bacteria failed to produce functional insulin protein. When cDNA was used instead, the bacteria successfully produced functional insulin.

(a) Why did the bacteria fail to produce functional insulin when genomic DNA was used as the source of the insulin gene? (1 mark)
(b) How is cDNA prepared from the insulin mRNA? Name the enzyme involved. (1 mark)
(c) Name the brand of the first recombinant human insulin approved for human use and the company that developed it. (1 mark)

A research team working on insulin production for diabetic patients used recombinant DNA technology to produce human insulin in bacteria. They isolated the human insulin gene from a human pancreatic cell and inserted it into a bacterial plasmid. The bacteria were then cultured in large bioreactors to produce insulin on an industrial scale.

(a) Name the type of insulin produced by this method and state ONE advantage it has over insulin extracted from animals. (1 mark)

(b) Draw a neat, labelled diagram of a stirred-tank bioreactor used for industrial production of recombinant insulin, showing at least FOUR labelled parts. (2 marks)

(c) Human insulin consists of two polypeptide chains A and B. Explain how scientists overcame the challenge of producing functional human insulin in bacteria, given that the insulin gene produces a precursor called pro-insulin. (2 marks)

A research team isolated a gene encoding human insulin and wanted to express it in Escherichia coli to produce recombinant human insulin. They used the restriction enzyme EcoRI to cut both the human insulin gene and the cloning vector pBR322. The recombinant plasmid was then introduced into E. coli cells.

(a) Draw a labelled diagram of the cloning vector pBR322, showing all essential features. Indicate clearly where the foreign gene (insulin gene) was inserted and how you would identify transformed colonies carrying the recombinant plasmid. (3 marks)

(b) The researchers found that when they directly cloned the human insulin gene (isolated from genomic DNA) into E. coli, no functional insulin protein was produced, even though the gene was successfully inserted. Suggest TWO reasons for this failure and explain how cDNA technology overcomes these problems. (2 marks)

Describe the steps involved in the production of a recombinant DNA molecule. Draw a labelled diagram showing the action of a restriction endonuclease on a DNA molecule with a palindromic sequence, resulting in sticky ends.