Unit-IX Biotechnology and its Applications
MCQs, Short Notes, Summary Notes, Flow Charts and Mnemonics, Rapid-Fire Revision for Board Exams and NEET, and CUET.
Chapter-11: Biotechnology - Principles and Processes
Genetic Engineering (Recombinant DNA Technology).
Chapter-12: Biotechnology and its Applications
Application of biotechnology in health and agriculture: Human insulin and vaccine production, stem cell technology, gene therapy; genetically modified organisms - Bt crops; transgenic animals;biosafety issues, biopiracy and patents.
📌 Practice these questions for best result.
🚀 Rapid-Fire Revision
✍️ First recombinant DNA molecule? – Paul Berg, 1972.
✍️ First genetically modified (GM) plant? – Tobacco.
✍️ First GM crop in India? – Bt cotton.
✍️ First recombinant drug approved for humans? – Insulin (Humulin).
✍️ First clinical gene therapy? – ADA deficiency (1990, USA).
✍️ Enzymes cutting DNA at specific sites? – Restriction endonucleases.
✍️ Sequence recognized by restriction enzyme? – Palindromic sequence.
✍️ Enzyme sealing DNA fragments? – DNA ligase.
✍️ Enzyme synthesizing cDNA? – Reverse transcriptase.
✍️ Enzyme used in PCR? – Taq DNA polymerase.
✍️ Steps of PCR? – Denaturation → Annealing → Extension.
✍️ Visualization dye for DNA in gel? – Ethidium bromide (under UV).
✍️ Common cloning host? – E. coli.
✍️ Most popular plasmid vector? – pBR322.
✍️ Method of DNA delivery with gene gun? – Biolistics.
✍️ Organism providing Bt gene (cry gene)? – Bacillus thuringiensis.
✍️ Bt toxin activates in insect due to? – Alkaline gut pH.
✍️ Golden rice is rich in? – Vitamin A (β-carotene).
✍️ Flavr Savr tomato – purpose? – Delayed ripening.
✍️ RNA interference (RNAi) silences what? – Specific mRNA.
✍️ Sheep transgenic for treating emphysema produces? – α1-antitrypsin.
✍️ Stem cells are promising for? – Regenerative medicine.
✍️ Recombinant vaccine developed for? – Hepatitis B.
✍️ Ex vivo gene therapy means? – Correcting cells outside body, reintroducing.
✍️ In vivo gene therapy means? – Direct delivery of genes into patient.
✍️ Biopiracy refers to? – Exploiting biodiversity without permission.
✍️ Examples of Indian biopiracy cases? – Turmeric, Neem, Basmati rice.
✍️ Law protecting biodiversity in India? – Biological Diversity Act, 2002.
✍️ Protocol for international biosafety? – Cartagena Protocol on Biosafety.
✍️ Major GM biosafety concerns? – Allergenicity, gene transfer, ecological imbalance.
✍️ 1 Mark Questions
Who constructed the first recombinant DNA molecule?
Answer: Paul Berg in 1972.
Why are restriction enzymes called molecular scissors?
Answer: Because they cut DNA at specific sites.
Name the recognition sequence for many restriction enzymes.
Answer: Palindromic sequence.
Which enzyme joins foreign DNA with vector DNA?
Answer: DNA ligase.
Name the most commonly used bacterial host in genetic engineering.
Answer: Escherichia coli.
What is the selectable marker in pBR322?
Answer: Antibiotic resistance genes (ampR, tetR).
Give an example of a cloning vector used in plants.
Answer: Ti plasmid of Agrobacterium tumefaciens.
Name the enzyme used in PCR.
Answer: Taq DNA polymerase.
What is cDNA?
Answer: DNA complementary to mRNA, synthesized by reverse transcriptase.
Which fluorescent dye is used to stain DNA in electrophoresis?
Answer: Ethidium bromide.
Name the bacterium from which Bt toxin gene is obtained.
Answer: Bacillus thuringiensis.
Which was the first hormone produced by recombinant DNA technology?
Answer: Insulin (Humulin).
Which disease was first treated by gene therapy?
Answer: ADA deficiency (Adenosine deaminase deficiency).
Golden rice is enriched with which vitamin?
Answer: Vitamin A (β-carotene).
Name one transgenic animal that produces α1-antitrypsin.
Answer: Transgenic sheep.
✍️ 2 Mark Questions
Differentiate between ex vivo and in vivo gene therapy.
Answer: Ex vivo: Cells are corrected outside the body and reintroduced.
In vivo: Therapeutic gene is directly introduced into patient’s body.
Why are plasmids used as vectors in rDNA technology?
Answer: They replicate independently, carry selectable markers, and have cloning sites.
What is the role of CaCl₂ in bacterial transformation?
Answer: It makes the bacterial cell membrane permeable to allow DNA entry.
Give any two features of pBR322 plasmid.
Answer: Ori site, selectable markers (ampR, tetR), multiple cloning sites.
What are the steps of PCR?
Answer: Denaturation, Annealing, Extension.
What are biofactories? Give an example.
Answer: Transgenic animals producing therapeutic proteins, e.g., sheep producing α1-antitrypsin.
Why is RNA interference (RNAi) useful in agriculture?
Answer: It silences specific genes of pests/pathogens, protecting crops from damage.
What is biopiracy? Give one example.
Answer: Exploitation of biodiversity/traditional knowledge without permission, e.g., patenting of Indian Basmati rice.
List two main biosafety concerns related to GMOs.
Answer: Allergenicity of food products, gene transfer to wild relatives.
Name two pest-resistant crops developed by biotechnology.
Answer: Bt cotton, Bt brinjal.
Differentiate between humulin and animal insulin.
Answer: Humulin: Produced by rDNA technology, identical to human insulin, no allergy risk.
Animal insulin: Derived from pig/cattle pancreas, could cause allergies.
What are the applications of stem cell technology?
Answer: Regenerative medicine: treatment of Parkinson's, diabetes, organ repair.
Why was Flavr Savr tomato developed?
Answer: To delay fruit ripening and spoilage using antisense RNA technology.
Explain why Bt toxin does not kill the bacterium producing it.
Answer: It is produced as an inactive protoxin in bacteria, activated only in alkaline insect gut.
State one role of biotechnology in the field of environment.
Answer: Development of bio-remediation organisms for cleaning oil spills, wastewater.
✍️ 5 Mark Questions
1. Describe the steps involved in the process of recombinant DNA technology.
Answer: The main steps are:
- Isolation of DNA from source organism.
- Cutting of DNA with restriction endonucleases at specific sites.
- Amplification of gene using PCR (Denaturation → Annealing → Extension).
- Insertion of gene into vector with DNA ligase.
- Introduction into host cells by transformation (CaCl₂, electroporation, biolistics, microinjection).
- Selection and screening of recombinants using selectable markers or blue/white selection.
- Multiplication and expression of gene in host leading to production of desired protein.
2. Explain the principle and steps of Polymerase Chain Reaction (PCR).
Answer:
Principle: Enzymatic amplification of a specific DNA sequence in vitro using thermostable DNA polymerase.
Steps:
- Denaturation: Heat (94–98°C) separates DNA strands.
- Annealing: Primers attach to complementary sequences (~55°C).
- Extension: Taq polymerase adds nucleotides to primers (~72°C).
This cycle is repeated ~30 times to obtain millions of DNA copies.
3. What are the desirable features of a cloning vector? Explain with example of pBR322.
Answer:
Desirable features:
- Origin of replication (Ori) – ensures replication.
- Selectable markers – antibiotic resistance genes (ampR, tetR) help identify recombinants.
- Multiple cloning sites (MCS) – sites for REs to insert foreign DNA.
- Small size – easy manipulation.
- High copy number – amplifies DNA efficiently.
pBR322 contains Ori, ampR, tetR, and unique restriction sites → widely used in E. coli.
4. Explain the process of producing human insulin using recombinant DNA technology.
Answer:
- The human insulin gene has two chains: A and B.
- Scientists (Eli Lilly, 1983) isolated and synthesized DNA sequences for both chains.
- Inserted into E. coli plasmids separately.
- Each chain produced in bacteria, isolated, purified.
- Chains were combined through disulphide bonds to form functional Humulin.
- Approved as the first recombinant therapeutic protein.
5. What is gene therapy? Explain the treatment process for ADA deficiency.
Answer:
The technique to treat diseases by correcting defective genes.
First used for ADA deficiency (Adenosine deaminase).
Process:
- Functional ADA gene is cloned from healthy individual.
- Introduced into patient’s lymphocytes (ex vivo).
- Genetically corrected lymphocytes are reintroduced into the patient.
- Provides functional ADA enzyme for immune function.
Limitation: periodic reinfusion needed; future: permanent gene insertion in stem cells.
6. Explain Bt cotton production. How does Bt toxin confer resistance to insects?
Answer:
Bt crops developed using cry genes from Bacillus thuringiensis.
- Cry IAc, Cry IIAb genes inserted into cotton genome.
- Bt protein is inactive protoxin in plant tissue.
- In insect gut (alkaline pH), protoxin → activated toxin.
- Activated toxin binds to gut epithelium → creates pores → insect dies.
- Provides natural insect resistance without chemical pesticide use.
7. What are transgenic animals? Mention any five applications.
Answer:
Animals into which foreign genes (from other species) are introduced.
Applications:
- Produce human proteins (e.g., α1-antitrypsin in sheep).
- Study of gene regulation and development.
- Create models for human diseases (cancer, diabetes).
- Produce vaccines and antibodies.
- Improve livestock for milk, meat, and wool quality.
8. Mention the benefits and risks of genetically modified (GM) crops.
Answer:
Benefits:
- Insect resistance (Bt cotton).
- Herbicide tolerance.
- Nutritional enhancement (Golden rice).
- Reduced pesticide use.
- Increased shelf life (Flavr Savr tomato).
Risks:
- Allergenicity in humans.
- Gene escape into wild relatives.
- Development of resistant pests.
- Loss of biodiversity.
- Ethical concerns.
9. What is RNA interference (RNAi)? How is it used in crop protection?
Answer:
RNAi: Method of silencing a gene post-transcription by using double-stranded RNA (dsRNA) which degrades target mRNA.
Insects/nematodes → dsRNA introduced against vital genes → mRNA degraded → no proper protein → pest dies.
Example: RNAi used in tobacco to prevent nematode infectation.
10. What is biopiracy? Explain with examples and mention laws that prevent it in India.
Answer:
Use/exploitation of bioresources and traditional knowledge without proper authorization/compensation is biopiracy.
Examples:
Patenting of Indian Basmati rice by US companies.
US patents on Neem products and turmeric healing property.
Prevention: Biological Diversity Act (India, 2002) ensures fair access and benefit sharing; Cartagena Protocol on Biosafety ensures safe use of GMOs.