Biotechnology and Its ApplicationsNEET Botany · Class 12 · NCERT Chapter 8

Biotechnology and Its Applications contributes 3 to 4 questions per NEET paper. High-yield topics include: Bt toxin mechanism (cry genes, protoxin to toxin in alkaline gut, pore formation in bollworm), Bt cotton (cry1Ac, cry2Ab), RNA interference with Meloidogyne incognita and tobacco, recombinant insulin (humulin, Eli Lilly 1982, A and B chains in E. coli), ADA gene therapy (Ashanthi De Silva, 1990, retroviral vector), ELISA principle (antigen-antibody, enzyme-conjugated secondary antibody, colour substrate), and biopiracy examples.

Bt toxin refers to Cry proteins (Cry toxins) produced by the soil bacterium Bacillus thuringiensis. The toxin is produced as an inactive protoxin (proform) stored in bacterial spores. In the highly alkaline gut of insect larvae (e.g., cotton bollworm), the protoxin is activated (converted to active toxin). The active toxin binds specifically to receptor proteins on the epithelial cells lining the larval gut. This binding creates pores (holes) in the cell membrane, causing the cells to swell and lyse (burst). The gut becomes leaky, the larva stops feeding and dies. Bt toxin is safe for mammals: in humans and animals, the stomach is acidic, so the protoxin is not activated, and mammals lack the specific gut receptor proteins.

Bt cotton expresses two cry genes: (1) cry1Ac — controls American bollworm (Helicoverpa armigera), a Lepidoptera pest. (2) cry2Ab — additional Lepidoptera coverage. Bt corn expresses cry1Ab (controls corn borer, Lepidoptera). The choice of cry gene depends on the target pest order: cry1 and cry2 genes are effective against Lepidoptera; cry3 genes are effective against Coleoptera (beetles).

RNA interference (RNAi) is a cellular mechanism for post-transcriptional gene silencing. When double-stranded RNA (dsRNA) complementary to a specific mRNA is present in a cell, the RISC (RNA-induced silencing complex) is activated. RISC unwinds the dsRNA, uses one strand as a guide, and degrades the complementary mRNA, silencing the gene. Application: Transgenic tobacco was developed to resist the nematode Meloidogyne incognita (root-knot nematode). The transgenic plant produces dsRNA specific to a gene essential for nematode survival. When nematodes feed on the transgenic roots, the dsRNA enters nematode cells, triggers RNAi, silences the target nematode gene, and prevents the parasite from completing its life cycle in the plant root.

Before recombinant DNA technology: insulin was extracted from pig and cattle pancreas. Animal insulin differs from human insulin in a few amino acids and caused allergic reactions in some patients. Recombinant human insulin (brand name humulin, developed by Eli Lilly in 1982): (1) The gene for human insulin A chain was synthesised and cloned into E. coli. (2) The gene for insulin B chain was similarly cloned into E. coli. (3) Each chain is produced separately in E. coli under the control of a promoter. (4) The chains are extracted, purified, and combined. Disulfide (S-S) bonds form between the chains to produce functional insulin identical to human pancreatic insulin. It causes no allergic reactions and is available in unlimited quantities.

Adenosine deaminase (ADA) deficiency is a genetic disorder caused by deletion or mutation in the ADA gene. ADA is required to convert adenosine to inosine in the purine salvage pathway. Without ADA, toxic deoxyadenosine accumulates in T lymphocytes, destroying them and causing severe combined immunodeficiency (SCID). Treatment by gene therapy: (1) Lymphocytes are isolated from the patient's blood. (2) A functional ADA gene is inserted into a retroviral vector. (3) The retroviral vector infects the lymphocytes, integrating the ADA gene into the lymphocyte genome. (4) The genetically corrected lymphocytes are returned to the patient. The lymphocytes can now produce ADA and survive. The first clinical trial was performed on Ashanthi De Silva in 1990. This is not a permanent cure — the patient needs periodic infusions of corrected lymphocytes because lymphocytes are not stem cells and are not self-renewing. A permanent cure would require introduction of ADA gene into bone marrow stem cells.

ELISA (Enzyme Linked Immunosorbent Assay) is a diagnostic technique that detects antigens or antibodies in a sample using enzyme-labelled antibodies. Basic principle: (1) The sample (containing antigen) is added to a plate coated with a primary antibody (or antigen). (2) The primary antibody binds the specific antigen. (3) An enzyme-conjugated secondary antibody (anti-antibody) is added — it binds to the primary antibody. (4) A substrate for the enzyme is added. The enzyme converts the colourless substrate to a coloured product. The colour intensity is proportional to the amount of antigen present. Uses: HIV diagnosis (detects anti-HIV antibodies or HIV proteins), pregnancy tests (detects HCG hormone), hepatitis B detection, allergen testing, drug monitoring.