Tissue culture and somatic hybridisation
Thousands of identical banana plants can be grown from tiny pieces of one plant in a laboratory, free of disease.
Tissue culture is growing whole plants from an explant (any plant part) in a sterile nutrient medium. This is possible because plant cells are totipotent. Producing thousands of plants this way is micropropagation, and the plants are identical somaclones. Somatic hybridisation fuses protoplasts of two varieties to form a hybrid.
In the figure : Plant growth regulators (auxins and cytokinins) guide the formation of shoots and roots.
In the figure : Pomato (potato + tomato) was produced but lacked the desired characteristics.
Rule 1: Micropropagation and virus-free plants — The meristem (apical and axillary) of an infected plant is free of virus, so culturing it gives virus-free plants (banana, sugarcane, potato). Tomato, banana and apple are propagated commercially by micropropagation.
- Easy : Meristem is virus-free.
- Medium : Somaclones are genetically identical.
- Exam-type : Q: Why is meristem culture used to get virus-free plants? A: The meristem tip is free from virus.
Rule 2: Protoplast fusion — Cell walls are digested to get naked protoplasts. Protoplasts of two different plants are fused to form a hybrid protoplast, which is grown into a new plant. These hybrids are called somatic hybrids.
- Easy : Protoplast = cell without a wall.
- Medium : Somatic hybrid combines both parents.
- Exam-type : Q: What is a protoplast? A: A plant cell with its cell wall removed.
Totipotency = ability of a cell to grow into a whole plant. Medium: sucrose (carbon source), salts, vitamins, amino acids, auxin, cytokinin.
Thinking tissue culture plants are variable. They are clones of the parent plant.
Memory clue: T-M-S — Totipotency → Micropropagation → Somaclones.
Spot the difference
| Point | Micropropagation | Somatic hybridisation |
|---|---|---|
| Starting material | Explant | Two protoplasts |
| Result | Clones | Hybrid plant |
| Example | Banana plantlets | Pomato |
Practice questions
- 1What is totipotency?
- 2What is an explant?
- 3What are somaclones?
- 4Name two growth regulators used in a culture medium.
- 5What is somatic hybridisation?
- 6Name the hybrid of potato and tomato.
- 7Why is the plant cell wall digested before fusion?
- 8Why is tissue culture useful for farmers?
Show answers (try them yourself first)
- 1The ability of a plant cell to grow into a complete plant.
- 2A plant part used for starting a culture.
- 3Genetically identical plants from tissue culture.
- 4Auxin and cytokinin.
- 5Fusion of protoplasts from different plants to form a hybrid.
- 6Pomato.
- 7The wall blocks fusion; naked protoplasts can fuse.
- 8Large numbers of healthy, identical plants are produced quickly.
Genetically modified crops: Bt cotton, RNAi and golden rice
A cotton farmer who sprays less pesticide because the plant makes its own insect-killing protein saves money and protects the environment.
Genetically modified organisms (GMOs) have genes altered by manipulation. GM crops are tolerant to stress, need less pesticide, have a longer shelf life, and have better food value (golden rice is rich in vitamin A). Bt cotton carries a cry gene from Bacillus thuringiensis that produces an insecticidal protein.
In the figure : The toxin is inactive in the bacterium and in the plant but becomes active in the alkaline gut of the insect.
Rule 1: Bt toxin — The genes cryIAc and cryIIAb control the cotton bollworms; cryIAb controls the corn borer. The protein is specific to groups of insects, so harmless insects are not harmed. The toxin forms pores in the midgut.
- Easy : Bt cotton resists bollworms.
- Medium : Bt toxin is inactive as protoxin.
- Exam-type : Q: Why does the Bt toxin not kill the bacterium? A: It is present as an inactive protoxin in the bacterium.
Rule 2: RNA interference (RNAi) — RNAi silences a specific mRNA by a complementary double-stranded RNA. The nematode Meloidogyne incognita infects tobacco roots and reduces the yield. A transgenic tobacco plant was made to produce complementary dsRNA, which silenced the nematode's gene and the parasite could not survive.
- Easy : RNAi silences genes.
- Medium : It is based on complementary RNA.
- Exam-type : Q: What did RNAi do in tobacco roots? A: It silenced a gene of the parasite, protecting the plant from the nematode.
Golden rice: vitamin A. Bt: Bacillus thuringiensis. RNAi: gene silencing by dsRNA that is complementary to the mRNA.
Writing that the Bt toxin kills all insects. It is group-specific.
Memory clue: Bt → Bacillus thuringiensis → Bollworm tamer — Bt cotton protects against bollworms.
Spot the difference
| Point | Bt cotton | RNAi plant |
|---|---|---|
| Gene source | Bacillus thuringiensis | Complementary dsRNA |
| Target | Cotton bollworm | Nematode (Meloidogyne) |
| Mechanism | Toxin kills larvae | Silences parasite mRNA |
Practice questions
- 1What does GMO stand for?
- 2Name the gene for the Bt toxin in cotton.
- 3What is golden rice rich in?
- 4What is RNAi?
- 5Name the nematode that infects tobacco roots.
- 6Give one advantage of GM crops.
- 7Why is the alkaline gut necessary for Bt toxin action?
- 8How does a transgenic tobacco plant resist the nematode?
Show answers (try them yourself first)
- 1Genetically modified organism.
- 2cry gene (e.g. cryIAc).
- 3Vitamin A.
- 4Silencing of a specific mRNA by a complementary double-stranded RNA.
- 5Meloidogyne incognita.
- 6Any: stress tolerance, less pesticide use, longer shelf life, higher nutritive value.
- 7It dissolves the crystals and makes the protoxin active.
- 8It makes dsRNA that silences a nematode gene essential for the parasite.
Biotechnology in medicine: insulin, gene therapy and diagnosis
Diabetics once used insulin from cattle and pigs. Today human insulin is made in bacteria that carry the human gene.
Recombinant therapeutics are safer because they are identical to the human protein. In 1983 Eli Lilly made human insulin by making DNA for chains A and B and inserting them in E. coli plasmids; the chains were produced separately and joined by disulphide bonds. Gene therapy corrects a gene defect; the first clinical gene therapy (1990) was for ADA deficiency.
In the figure : In humans insulin is made as pro-insulin; the C peptide is removed to make the mature hormone.
In the figure : Both give early detection before symptoms are clear.
Rule 1: Gene therapy — ADA is needed for a healthy immune system; its absence makes a child lack immunity. Treatment by enzyme replacement is not curative. In gene therapy, lymphocytes from the patient are grown in culture, a functional ADA cDNA is introduced using a retroviral vector, and the cells are returned. These cells are not immortal, so periodic infusion is needed unless gene-introduced cells are from early embryonic cells.
- Easy : ADA deficiency affects immunity.
- Medium : A retrovirus carries the ADA cDNA.
- Exam-type : Q: What is gene therapy? A: A method to correct a gene defect by inserting a normal gene.
Rule 2: Molecular diagnosis — PCR detects very low levels of pathogen DNA (for example HIV) and mutations in cancer. A radioactive single-stranded DNA probe hybridises to its complementary DNA; the clone with the mutated gene does not show up on the autoradiogram. ELISA works on antigen–antibody interaction.
- Easy : ELISA detects antibodies or antigens.
- Medium : Probes find complementary sequences.
- Exam-type : Q: Why is PCR valuable in detecting HIV? A: It can detect very low amounts of viral nucleic acid early.
Recombinant human insulin does not cause allergy as animal insulin sometimes does. ADA gene therapy: first done in 1990 on a 4-year-old girl.
Thinking insulin chains A and B are made as a single chain. They are made separately and joined by disulphide bonds.
Memory clue: A + B → I — Chain A + chain B, joined by S–S bonds = Insulin.
Spot the difference
| Point | Gene therapy | Enzyme replacement |
|---|---|---|
| Method | Insert a normal gene | Give the enzyme |
| Duration | Longer-lasting | Needs repeat dosing |
| Cure | Possible | Not curative |
Practice questions
- 1Which company first made human insulin in E. coli?
- 2How many chains does mature insulin have?
- 3What bonds join chains A and B?
- 4What is ADA?
- 5What is a probe?
- 6What is ELISA based on?
- 7Why is recombinant insulin better than animal insulin?
- 8Why are the corrected cells in gene therapy given repeatedly?
Show answers (try them yourself first)
- 1Eli Lilly (1983).
- 2Two (A and B).
- 3Disulphide bonds.
- 4Adenosine deaminase, an enzyme needed for immunity.
- 5A tagged single-stranded DNA or RNA that hybridises with its complementary sequence.
- 6Antigen–antibody interaction.
- 7It is identical to human insulin and does not cause allergy or immune reaction.
- 8Lymphocytes are not immortal, so the treatment must be repeated.
Transgenic animals and ethical issues
A mouse that carries a human disease gene helps scientists test new medicines without testing on people first.
Transgenic animals carry and express a foreign gene; over 95 per cent are mice. Uses: study of gene function, disease models (cancer, cystic fibrosis, Alzheimer's), biological products (α-1-antitrypsin; the transgenic cow Rosie, 1997), vaccine safety and chemical safety testing. Biopiracy is the use of bio-resources by companies without proper authorisation.
In the figure : Transgenic animals replace tests on humans and larger animals.
Rule 1: Applications — Rosie produced human α-lactalbumin-rich milk (2.4 g/litre), a better balanced food for human babies. α-1-antitrypsin is used to treat emphysema. Transgenic mice are used to test polio vaccine safety and could replace monkeys.
- Easy : Rosie was the first transgenic cow (1997).
- Medium : Transgenic mice replace monkeys in polio vaccine tests.
- Exam-type : Q: Which protein did Rosie's milk contain? A: Human α-lactalbumin.
Rule 2: Ethics, GEAC, patents and biopiracy — GEAC (Genetic Engineering Approval Committee) decides the safety of GM research and putting GM organisms into public use. Patents on biological materials (for example Basmati rice, turmeric, neem) led to concern. India has laws such as the Indian Patents Bill, and Biopiracy needs to be prevented because traditional knowledge belongs to its community.
- Easy : GEAC regulates GM research.
- Medium : Basmati and turmeric patents raised concern.
- Exam-type : Q: What is biopiracy? A: Use of biological resources and traditional knowledge by organisations without proper authorisation or payment.
GEAC = Genetic Engineering Approval Committee. Over 95 per cent of transgenic animals are mice.
Thinking GM research needs no approval. In India GEAC must approve it.
Memory clue: GEAC → Gatekeeper — GEAC guards the safety of GM organisms.
Spot the difference
| Point | Transgenic animal | GMO plant |
|---|---|---|
| Example | Rosie the cow | Bt cotton |
| Purpose | Products, disease models | Pest resistance, nutrition |
| Common species | Mice | Cotton, rice |
Practice questions
- 1What is a transgenic animal?
- 2Which animal is used most often?
- 3Name the first transgenic cow.
- 4Expand GEAC.
- 5What is biopiracy?
- 6Name one human protein made in transgenic animals.
- 7Why are transgenic mice used in vaccine safety testing?
- 8Why should GM organisms be regulated?
Show answers (try them yourself first)
- 1An animal that has a foreign gene in its genome and expresses it.
- 2Mice (over 95 per cent).
- 3Rosie.
- 4Genetic Engineering Approval Committee.
- 5Unauthorised use of biological resources or traditional knowledge.
- 6α-1-antitrypsin (or α-lactalbumin).
- 7They can replace monkeys and give quicker results.
- 8To check their risks for health and the environment before release.