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State Board · Class 12 · Biology

Lesson 12: Biotechnology

7 concepts · 56 practice questions with answers · Free to read

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Biotechnology and tools of rDNA

Link to Std 9–10

Last lesson raised food by traditional methods; here we learn how changing genes directly gives new solutions.

From daily life

Curd to butter, yeast for bread — using organisms; now genes can be joined to create new traits.

Simple definition

Biotechnology = using organisms or their parts to make products/services. rDNA = new DNA made by joining DNA from two different sources.

rDNA toolsBiotechnology and tools of rDNAScissorsrestriction enzymeGlueDNA ligaseVectorcarrierHosthost cellBiotechnology: using organisms to make products/servicesCohen and Boyer (1973): first rDNA — Salmonella plasmidRestriction enzyme cuts DNA at a specific site; EcoRI → GAATTCPalindrome: same 5'→3' on both strands; sticky endsLigase joins fragments; vectors: plasmid (pBR322), phage, Ti plasmidHosts: E. coli, Agrobacterium, yeast, animal cellsExonuclease cuts from ends; endonuclease cuts inside

In the figure : Tools of rDNA.

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Biotechnology and tools of rDNA

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Rule 1 : Restriction enzymes and ligase — A restriction endonuclease cuts DNA at a specific sequence; the sequence is a palindrome — same 5'→3' on both strands. E.g. EcoRI: GAATTC; from E. coli RY13. Cuts leave sticky ends (easy to join). Exonuclease cuts from ends; endonuclease cuts inside. DNA ligase joins fragments. First rDNA: Cohen and Boyer, 1973 (a gene added to a Salmonella plasmid).

  • Easy : EcoRI: GAATTC.
  • Medium : Ligase = glue.
  • Exam-type (CET) : Restriction = scissors.

Rule 2 : Vectors and hosts — A vector carries a gene: plasmid (circular DNA; e.g. pBR322, ori + antibiotic marker), bacteriophage, Ti plasmid (Agrobacterium) for plants, retroviruses for animals. A good vector has an ori (replication), selectable marker, cloning site and small size. Hosts: E. coli, Agrobacterium, yeast, animal cells.

  • Easy : pBR322 = plasmid vector.
  • Medium : Ti = Agrobacterium.
  • Exam-type (CET) : Ori = replication.
Remember

Scissors + glue + vehicle + home.

Common mistake and CET trap

‘Ligase cuts DNA’ is wrong (joins). Trap: ‘palindrome on one strand only’ is wrong.

Shortcut

Memory clue : "Restriction rips; ligase links".

Spot the difference

ToolRole
Restriction enzymeCuts
DNA ligaseJoins
VectorCarries

Practice questions

Easy
  1. 1EcoRI site?
  2. 2Who made the first rDNA?
  3. 3Sticky ends are?
Medium
  1. 4Ligase function?
  2. 5pBR322 is?
  3. 6Plant vector?
Challenging
  1. 7Why is ori needed?
  2. 8Exonuclease cuts?
Show answers (try them yourself first)
  1. 1GAATTC.
  2. 2Cohen and Boyer, 1973.
  3. 3Single-stranded overhangs after cutting.
  4. 4Joins DNA fragments.
  5. 5A plasmid vector.
  6. 6Ti plasmid.
  7. 7To start replication.
  8. 8From the ends.

Gene cloning and PCR

From daily life

A photocopier makes many copies of a page — PCR makes millions of copies of a DNA piece.

Simple definition

Gene cloning = making many identical copies of a wanted gene. PCR = rapid in-vitro multiplication of a specific DNA region.

Cloning stepsSteps of gene cloningIsolateisolate DNACutcutJoinligateInserttransformSelectscreen1. Isolate DNA (lysozyme, cellulase, chitinase; RNase, protease)2. Cut with restriction enzyme; agarose gel electrophoresis3. Ligate into vector with ligase ⇒ recombinant DNA4. Into host: transformation, CaCl₂, microinjection, gene gun5. Selectable marker: antibiotic resistance; blue-white screeningPCR: denaturation 94°C → annealing 54°C → extension 72°CTaq polymerase (Thermus aquaticus) heat-stable; bioreactor follows

In the figure : Steps of gene cloning.

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Gene cloning and PCR

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Rule 1 : Cloning steps — (1) Isolate DNA: break cells — lysozyme (bacteria), cellulase (plants), chitinase (fungi); remove RNA/proteins with RNase and protease; precipitate DNA with ethanol. (2) Cut: restriction enzyme; fragments sorted by size on agarose gel electrophoresis; DNA is negative ⇒ moves to anode; seen with ethidium bromide + UV. (3) Join: ligase ⇒ recombinant DNA. (4) Insert in host: transformation (CaCl₂, heat shock), microinjection, gene gun. (5) Select: antibiotic marker, blue-white screening (insertion in lacZ ⇒ white colony).

  • Easy : DNA → anode.
  • Medium : Lysozyme = bacteria.
  • Exam-type (CET) : White colony = recombinant.

Rule 2 : PCR — Polymerase Chain Reaction (Kary Mullis, 1983). Components: template DNA, two primers, Taq polymerase (Thermus aquaticus, heat-stable), dNTPs. Cycle: denaturation 94°C (strands separate) → annealing ~54°C (primers bind) → extension 72°C (new strands). Copies double each cycle; after n cycles 2ⁿ. Uses: disease diagnosis (HIV, TB), forensics, gene cloning. Next: large-scale production in a bioreactor (stirred-tank, sparged).

  • Easy : 94 → 54 → 72°C.
  • Medium : Taq = heat-stable.
  • Exam-type (CET) : 2ⁿ copies.
Remember

D-A-E: denature, anneal, extend.

Common mistake and CET trap

‘PCR uses ligase’ is wrong (Taq). Trap: ‘DNA moves to cathode’ is wrong (anode).

Shortcut

Memory clue : "DNA is negative → moves to anode".

Spot the difference

StepTemperature
Denaturation94°C
Annealing~54°C
Extension72°C

Practice questions

Easy
  1. 1Inventor of PCR?
  2. 2Source of Taq?
  3. 3Denaturation temperature?
Medium
  1. 4Copies after 5 cycles?
  2. 5DNA moves toward?
  3. 6Gene gun delivers?
Challenging
  1. 7White colony means?
  2. 8Enzyme to lyse plant cells?
Show answers (try them yourself first)
  1. 1Kary Mullis.
  2. 2Thermus aquaticus.
  3. 394°C.
  4. 432 (2⁵).
  5. 5Anode.
  6. 6DNA-coated particles into cells.
  7. 7Recombinant.
  8. 8Cellulase.

Medical applications

From daily life

Insulin for diabetics once came from animals; now bacteria make it.

Simple definition

A recombinant protein = protein made through rDNA technology. Gene therapy = giving a healthy gene to replace a faulty one.

Medical usesMedical applicationsInsulindiabetesVaccinessafe vaccinesGene therapygene correctionInsulin: A and B chains; C-peptide removed; Eli Lilly 1983 (Humulin)Pro-insulin → mature insulin; chains made separately in E. coliRecombinant vaccine: hepatitis B (yeast), safe, no infectionGene therapy: ADA deficiency (SCID); 1990; healthy gene replaces faulty oneMolecular diagnosis: PCR, ELISA, DNA probe — early detectionGenetic counselling, DNA fingerprinting, stem cellsInterferon, growth hormone, clotting factors — recombinant

In the figure : Insulin, vaccines, gene therapy.

Rule 1 : Insulin and vaccines — Insulin: 51 amino acids; two chains A (21) and B (30) joined by disulphide bonds. Animal insulin caused allergy; so Eli Lilly in 1983 made genes for the A and B chains separately in E. coli and joined them (Humulin). In pro-insulin the middle C-peptide is removed to give mature insulin. A recombinant vaccine, e.g. hepatitis B (surface antigen in yeast) — immunity without infection. Others: growth hormone, interferon, clotting factor VIII.

  • Easy : A 21 + B 30.
  • Medium : C-peptide is removed.
  • Exam-type (CET) : Humulin 1983.

Rule 2 : Gene therapy and diagnosis — Gene therapy: first in 1990, for ADA deficiency (SCID) — no adenosine deaminase, weak immunity; healthy gene inserted into the patient's lymphocytes; given to stem cells it becomes permanent. Diagnosis: PCR (early HIV, TB), ELISA, DNA probe (single-stranded DNA with radioactive/fluorescent tag; binds a complementary sequence). DNA fingerprinting, genetic counselling.

  • Easy : ADA = SCID.
  • Medium : Probe = complementary sequence.
  • Exam-type (CET) : Stem cells = lasting.
Remember

Insulin = A + B (21 + 30).

Common mistake and CET trap

‘Insulin is one chain’ is wrong (two). Trap: ‘gene therapy is just a drug’ is wrong (gene).

Shortcut

Memory clue : "A 21 + B 30 = 51".

Spot the difference

ProductSource/host
HumulinE. coli
Hepatitis B vaccineYeast
Gene therapyADA gene

Practice questions

Easy
  1. 1Amino acids in insulin?
  2. 2Chain A has?
  3. 3Humulin year?
Medium
  1. 4C-peptide?
  2. 5Host for hepatitis B vaccine?
  3. 6ADA deficiency causes?
Challenging
  1. 7DNA probe is?
  2. 8ELISA is used for?
Show answers (try them yourself first)
  1. 151.
  2. 221 amino acids.
  3. 31983.
  4. 4Removed in maturation.
  5. 5Yeast.
  6. 6SCID.
  7. 7Tagged single-stranded DNA.
  8. 8Detecting antibody/antigen.

Transgenic plants

From daily life

Bollworm damaged cotton; Bt cotton reduced spraying.

Simple definition

Transgenic = organism carrying a gene from another species (GMO). A GM crop = genetically modified crop.

GM cropsTransgenic plantsBt cottonpest resistantGolden ricevitamin AFlavr Savrdelayed ripeningTransgenic = organism carrying a foreign gene (GMO)Bt cotton: cry gene; Cry protein makes holes in the larval gutCry1Ac — bollworm; Cry1Ab — corn borerGolden rice: β-carotene → vitamin A (prevents blindness)Flavr Savr tomato: pectinase blocked, longer shelf lifeRNA interference: tobacco against Meloidogyne (nematode)Agrobacterium tumefaciens: Ti plasmid vector; dicots

In the figure : Bt cotton, golden rice, Flavr Savr.

Rule 1 : Bt cotton — The Bacillus thuringiensis cry gene (Cry1Ac, Cry1Ab) is placed in cotton. The bacterium makes a protoxin that turns active in the larva's alkaline gut ⇒ holes in gut cells ⇒ larva dies. The toxin does not activate in humans or other animals. Use: protection from bollworm; less spraying. Vector: Agrobacterium Ti plasmid. India approved Bt cotton in 2002.

  • Easy : Cry = larval toxin.
  • Medium : Alkaline gut.
  • Exam-type (CET) : Ti plasmid = vector.

Rule 2 : Golden rice, Flavr Savr, RNAi — Golden rice: rice with β-carotene (vitamin A precursor); genes from daffodil and a bacterium; reduces vitamin A deficiency and night blindness. Flavr Savr tomato: blocking polygalacturonase (pectin-degrading) delays softening — longer shelf life (1994). RNA interference (RNAi): RNA complementary to a gene of Meloidogyne incognita (nematode) ⇒ silences its mRNA ⇒ protects tobacco roots. Benefits in plants: higher yield, nutrition, less pesticide.

  • Easy : Golden = β-carotene.
  • Medium : Flavr Savr = long shelf life.
  • Exam-type (CET) : RNAi silences mRNA.
Remember

Bt: poison for the larva, not for us.

Common mistake and CET trap

‘Bt toxin is harmful to all animals’ is wrong. Trap: ‘golden rice gives iron’ is wrong (vitamin A).

Shortcut

Memory clue : "Golden rice = carrot-like β-carotene".

Spot the difference

GM cropTrait
Bt cottonPest-resistant
Golden riceVitamin A
Flavr SavrLonger shelf life

Practice questions

Easy
  1. 1Bt stands for?
  2. 2Cry protein is?
  3. 3Larval gut is?
Medium
  1. 4Golden rice has?
  2. 5Flavr Savr is a?
  3. 6RNAi works by?
Challenging
  1. 7Agrobacterium gives?
  2. 8Transgenic means?
Show answers (try them yourself first)
  1. 1Bacillus thuringiensis.
  2. 2Insecticidal toxin.
  3. 3Alkaline.
  4. 4β-carotene.
  5. 5Delayed-ripening tomato.
  6. 6Silencing mRNA.
  7. 7Ti plasmid vector.
  8. 8Carrying a foreign gene.

Transgenic animals and clones

From daily life

Milk of a goat that yields a medicinal protein — an animal as a tiny factory.

Simple definition

A transgenic animal = animal carrying a foreign gene. A clone = genetically identical animal made from one cell.

Transgenic animalsTransgenic animalsMousemodelRosie cowhuman milk proteinFish / Goatdrugs/fishAims: disease study, vaccine safety tests, drugs, nutritionTransgenic mouse: polio vaccine tests; cancer/diabetes modelsRosie (1997) cow: milk with α-lactalbuminTransgenic fish: fast growth; goat: antithrombinDolly (1996) sheep: a clone (nuclear transfer), not transgenicEthics: animal welfare, biosafety, ecosystem risksKnock-out mouse: to learn a gene's function

In the figure : Transgenic animals.

Rule 1 : Uses — (1) Disease models: transgenic mice for cancer, diabetes, Alzheimer's. (2) Vaccine and drug-safety tests: e.g. polio vaccine tests — mice instead of monkeys. (3) Biological products: Rosie cow (1997) — milk with human α-lactalbumin; goat milk — antithrombin. (4) Chemical safety testing. (5) Nutrition: transgenic fish — fast growth. Knock-out mice: switching off a gene shows its function. Dolly (1996) sheep: cloned by nuclear transfer — not transgenic.

  • Easy : Rosie = α-lactalbumin.
  • Medium : Dolly = clone.
  • Exam-type (CET) : Knock-out = gene off.

Rule 2 : Ethics — Questions: animal suffering (animal welfare), gene flow to nature, ecosystem impact, human health, religious/ethical objections. Hence regulatory committees and laws; permission and monitoring for experiments; alternatives (computer models, cell culture); the ‘3R’ principle — Replace, Reduce, Refine. Responsible use is essential.

  • Easy : 3R: Replace, Reduce, Refine.
  • Medium : Regulatory committee.
  • Exam-type (CET) : Gene-flow risk.
Remember

Dolly = duplicate; Rosie = rich milk.

Common mistake and CET trap

‘Dolly is transgenic’ is wrong (clone). Trap: ‘no rules for transgenic animals’ is wrong.

Shortcut

Memory clue : "Rosie → protein-rich milk".

Spot the difference

AnimalSpecialty
RosieHuman milk protein
DollyClone
Knock-out mouseGene silenced

Practice questions

Easy
  1. 1Who is Rosie?
  2. 2Rosie's milk contains?
  3. 3Dolly was a?
Medium
  1. 4Knock-out mouse?
  2. 5Transgenic fish show?
  3. 63R means?
Challenging
  1. 7One ethical concern?
  2. 8Transgenic mice used for?
Show answers (try them yourself first)
  1. 1Transgenic cow (1997).
  2. 2α-lactalbumin.
  3. 3Cloned sheep.
  4. 4Mouse with a gene switched off.
  5. 5Faster growth.
  6. 6Replace, Reduce, Refine.
  7. 7Animal welfare.
  8. 8Vaccine safety tests.

Biosafety and GEAC

From daily life

A new medicine needs trials and approval before sale — so do GM crops.

Simple definition

Biosafety = protecting health and environment from biotechnology products. GEAC = Genetic Engineering Appraisal Committee.

BiosafetyBiosafety, GEAC, patentsGEACapproval bodyBiopatentbio-patentBiopiracybio-theftGEAC: Genetic Engineering Appraisal Committee (India)Role: check safety of GM crops/products, approve public useConcerns: allergy, toxicity, ecosystem impact, gene flowBiopatent: rights over a biological product/processBiopiracy: using traditional knowledge/resources without consent/sharinge.g. basmati, turmeric, neem patent disputes; Indian patent lawBiological Diversity Act 2002: local benefit and protection

In the figure : Biosafety and GEAC.

Rule 1 : Concerns and GEAC — Concerns: allergy, toxicity, effects on other species (e.g. butterflies), gene flow (GM crop to weeds), spread of antibiotic resistance, biodiversity loss. In India GEAC (Ministry of Environment) checks safety of GM crops/products before public use and approves field trials. International: Cartagena Protocol (2000). Principle: balance benefit and risk.

  • Easy : GEAC = safety approval.
  • Medium : Gene-flow risk.
  • Exam-type (CET) : Cartagena 2000.

Rule 2 : Biopatent and biopiracy — A patent = exclusive right over an invention for a limited time. A biopatent = patent on a biological product/process. Biopiracy = using another nation's/community's biological resources or traditional knowledge without consent and benefit-sharing. Disputes: basmati (USA), turmeric (wound healing), neem (fungicide) — India showed prior traditional use and had patents revoked. India has the Biological Diversity Act, 2002; TKDL (Traditional Knowledge Digital Library).

  • Easy : Biopiracy = without consent.
  • Medium : TKDL = digital library.
  • Exam-type (CET) : Basmati, turmeric, neem.
Remember

GEAC guards; biopiracy borrows without asking.

Common mistake and CET trap

‘GEAC grants patents’ is wrong (safety approval). Trap: ‘biopiracy is legal trade’ is wrong.

Shortcut

Memory clue : "Basmati–turmeric–neem: B-T-N".

Spot the difference

TermMeaning
GEACSafety appraisal
BiopatentPatent on bio-product
BiopiracyUse without consent

Practice questions

Easy
  1. 1GEAC full form?
  2. 2Role of GEAC?
  3. 3Gene flow means?
Medium
  1. 4Biopatent is?
  2. 5Biopiracy is?
  3. 6A disputed patent?
Challenging
  1. 7India's Act?
  2. 8TKDL is?
Show answers (try them yourself first)
  1. 1Genetic Engineering Appraisal Committee.
  2. 2Check safety of GM products.
  3. 3GM gene spreading to weeds.
  4. 4Patent on a biological product.
  5. 5Using resources without consent.
  6. 6Basmati / turmeric / neem.
  7. 7Biological Diversity Act, 2002.
  8. 8Traditional Knowledge Digital Library.

Summary: tools, steps and uses

From daily life

In a kitchen: recipe, tools, cooking, serving — biotechnology also follows a sequence.

Simple definition

rDNA technology = cut + join + insert in vector + grow in host + apply. Responsible use matters.

SummarySummary: tools → steps → usesCut + JoinInsertApplyEcoRI + ligase ⇒ rDNApBR322 / Ti plasmid vector; E. coli / Agrobacterium hostPCR: Taq polymerase; 94 → 54 → 72°CInsulin (Humulin 1983), hep-B vaccine, gene therapy (ADA)Bt cotton, Golden rice, Flavr SavrGEAC = safety; biopiracy = unfair useDirection: benefit with responsibility

In the figure : Tools → steps → uses.

Rule 1 : Complete sequence — 1) Cut DNA with EcoRI (GAATTC). 2) Join into pBR322/Ti with ligase. 3) Put into E. coli/Agrobacterium (transformation). 4) Select by antibiotic marker / blue-white. 5) Multiply by PCR (94-54-72°C, Taq); large scale in a bioreactor. 6) Uses: insulin, vaccines, gene therapy; Bt cotton, golden rice; transgenic animals. 7) GEAC approval; consider biopatent and biopiracy.

  • Easy : Cut–join–insert–select.
  • Medium : PCR = copies.
  • Exam-type (CET) : GEAC = safety.

Rule 2 : Exam pairs — Cohen–Boyer 1973; Mullis PCR; Eli Lilly Humulin 1983; Taq = Thermus aquaticus; Cry1Ac = bollworm; golden rice = β-carotene; Flavr Savr = polygalacturonase blocked; Rosie = α-lactalbumin; Dolly = clone; GEAC = safety; basmati–turmeric–neem = biopiracy disputes.

  • Easy : Memorise the pairs.
  • Medium : Function over names.
  • Exam-type (CET) : Use with responsibility.
Remember

C-J-I-S-P-A: cut, join, insert, select, PCR, apply.

Common mistake and CET trap

Reversed: ligase joins; restriction cuts. Trap: ‘Dolly is transgenic’ is wrong.

Shortcut

Memory clue : "CJISPA".

Spot the difference

StageTool
CutEcoRI
JoinLigase
MultiplyPCR / Taq

Practice questions

Easy
  1. 1Year of first rDNA?
  2. 2Cutting enzyme?
  3. 3Joining enzyme?
Medium
  1. 4A host organism?
  2. 5PCR enzyme?
  3. 6Humulin is?
Challenging
  1. 7Bt cotton protects from?
  2. 8GEAC ensures?
Show answers (try them yourself first)
  1. 11973.
  2. 2Restriction endonuclease.
  3. 3DNA ligase.
  4. 4E. coli.
  5. 5Taq polymerase.
  6. 6Recombinant insulin.
  7. 7Bollworm.
  8. 8Safety of GM products.

Quick revision of the whole lesson

ConceptKey fact
rDNA toolsEcoRI GAATTC; ligase; pBR322
Cloning/PCR94-54-72°C; Taq; 2ⁿ
MedicineInsulin A21 + B30; ADA gene therapy
GM plantsBt cotton; golden rice; Flavr Savr
GM animalsRosie; knock-out; Dolly = clone
BiosafetyGEAC; biopiracy; Biodiversity Act 2002
SequenceCut → join → insert → select → apply

🎯 Most important points for the exam

  • Biotechnology and rDNA tools.
  • Gene cloning and PCR.
  • Medical applications.
  • Transgenic plants.
  • Transgenic animals and clones.
  • Biosafety, GEAC, patents.
  • Complete summary.
Study mantra

Understand the concept → draw the figure → memorise the key points → solve questions yourself.