100% Complete Notes
Paper I & Paper II

1.1 Structure of Atoms, Molecules and Chemical Bonds

• Atomic structure: nucleus (protons + neutrons) + electrons in orbitals; atomic number = protons; mass number = protons + neutrons
• Chemical bonds: Covalent (shared electrons — strongest; C–C, C–H, C–N, C–O in biomolecules); Ionic (electrostatic; NaCl; important in enzyme active sites); Hydrogen bonds (O–H···O, N–H···O; weaker individually but cumulative strength in DNA, proteins); Van der Waals (induced dipole; weakest; important in protein folding, enzyme-substrate complementarity)
• Hydrophobic interactions: non-polar groups cluster away from water; drives protein folding, membrane formation, lipid bilayer
• Covalent vs non-covalent bonds: biological systems use non-covalent bonds for reversible interactions (enzyme-substrate, antigen-antibody, receptor-ligand)

1.2 Biomolecules — Structure and Function

Carbohydrates

• Empirical formula (CH₂O)n; monosaccharides (glucose, fructose, galactose, ribose, deoxyribose); anomeric carbon → α and β forms; ring closure (Haworth projection)
• Disaccharides: Sucrose (α-glucose + β-fructose; 1→2 glycosidic bond — non-reducing); Lactose (β-galactose + glucose; 1→4 bond — reducing); Maltose (glucose + glucose; α-1,4 — reducing)
• Polysaccharides: Starch (amylose — unbranched α-1,4 links; amylopectin — α-1,4 + α-1,6 branches); Glycogen (highly branched, α-1,6 every 8–10 glucose — rapid mobilisation); Cellulose (β-1,4 — unbranched; structural; humans cannot digest — dietary fibre); Chitin (β-1,4 N-acetylglucosamine — fungal cell walls, arthropod exoskeleton)
• Glycoproteins and glycolipids: carbohydrate covalently attached to protein/lipid; cell recognition, blood group antigens (ABO — N-acetylgalactosamine, galactose on red cell surface)

Lipids

• Fatty acids: carboxyl head + hydrocarbon chain; saturated (no double bonds — palmitic, stearic; solid at RT); unsaturated (1+ double bonds — oleic [mono], linoleic, linolenic [poly]; cis configuration → kink → lower melting point)
• Triacylglycerols (triglycerides): glycerol + 3 fatty acids; energy storage (9 kcal/g); adipose tissue
• Phospholipids: glycerol + 2 fatty acids + phosphate head group; amphipathic → form bilayers; examples: phosphatidylcholine (lecithin), phosphatidylinositol (PIP₂ signalling), phosphatidylserine (cell death signal on outer leaflet)
• Sterols: fused 4-ring structure; Cholesterol (membrane fluidity regulator — stiffens at high temp, prevents solidification at low temp; precursor for steroid hormones, bile acids, vitamin D); Phytosterols (plant); Ergosterol (fungal)
• Sphingolipids: sphingosine backbone; sphingomyelin (myelin sheath); cerebrosides and gangliosides (neural function; ABO antigens on red cells are glycolipids)
• Isoprenoids/Terpenes: isoprene (C5) building blocks; monoterpenes (menthol), sesquiterpenes, diterpenes (Vitamin A, retinal), tetraterpenes (carotenoids — β-carotene, lycopene), polyterpenes (rubber, dolichol)

Proteins

• Amino acids: 20 standard; α-amino acid — NH₂, COOH, H, R group on α-carbon; L-configuration; zwitterion at physiological pH
• Classification by R group: non-polar aliphatic (Gly, Ala, Val, Leu, Ile, Pro, Met); aromatic (Phe, Trp, Tyr); polar uncharged (Ser, Thr, Cys, Asn, Gln); positively charged at pH 7 (Lys, Arg, His); negatively charged at pH 7 (Asp, Glu)
• Peptide bond: condensation between α-COOH and α-NH₂; planar; partial double bond character → no free rotation; cis or trans (trans preferred — less steric clash)
• Protein structure levels: Primary (amino acid sequence — dictates all higher levels); Secondary (local regular folding — α-helix: 3.6 residues/turn, H-bond i to i+4; β-sheet: parallel or antiparallel H-bonding between extended strands; β-turns; Ω-loops); Tertiary (overall 3D folding — held by hydrophobic core, H-bonds, ionic bonds, disulfide bridges); Quaternary (multiple subunits; haemoglobin = α₂β₂)
• Ramachandran plot: allowed φ and ψ (backbone dihedral angles); right-handed α-helix and β-sheets are in allowed regions; proline and glycine have unique constraints
• Protein folding: hydrophobic collapse drives folding; chaperones (Hsp70, GroEL/GroES) assist and prevent aggregation; prions — misfolded proteins that template misfolding of native proteins (PrPᶜ → PrPˢᶜ)

Nucleic Acids

• Nucleotide = nitrogenous base + pentose sugar + 1–3 phosphates; nucleoside = base + sugar only
• Purines: Adenine (A) and Guanine (G) — double ring; Pyrimidines: Cytosine (C), Thymine (T in DNA), Uracil (U in RNA) — single ring
• DNA: B-form most common (Watson-Crick 1953); right-handed double helix; antiparallel strands (5'→3' and 3'→5'); A–T (2 H-bonds), G–C (3 H-bonds); major and minor grooves; A-form (RNA duplexes, short RNA–DNA), Z-form (left-handed; GC-rich; role in gene regulation)
• RNA: single-stranded; U instead of T; ribose; mRNA (protein-coding), tRNA (adaptor — anticodon loop; aminoacyl-tRNA synthetase charges tRNA), rRNA (ribosome — structural + catalytic; 23S rRNA is the ribozyme that forms peptide bonds), snRNA (splicing), miRNA/siRNA (gene silencing), lncRNA (chromatin regulation)
• Conformation of tRNA: L-shaped 3D structure; CCA-3' at 3' end (amino acid attachment); anticodon loop at opposite end

Vitamins and Coenzymes

1.3 Stabilizing Interactions and Biophysical Chemistry

• pH and pKa: Henderson-Hasselbalch equation pH = pKa + log([A⁻]/[HA]); at pH = pKa, 50% ionised; physiological pH 7.4
• Buffers: resist pH change; bicarbonate buffer (blood — H₂CO₃/HCO₃⁻, pKa 6.1); phosphate buffer (intracellular — H₂PO₄⁻/HPO₄²⁻, pKa 6.8); protein buffer (histidine residues, pKa ~6)
• Thermodynamics: Gibbs free energy ΔG = ΔH – TΔS; spontaneous if ΔG < 0; equilibrium when ΔG = 0; ΔG° = –RT ln Keq; coupled reactions: exergonic (–ΔG) drives endergonic (+ΔG)
• ATP as energy currency: phosphoanhydride bonds (~7.3 kcal/mol each); ATP + H₂O → ADP + Pi (–7.3 kcal/mol); ATP → AMP + PPi (–7.3 kcal/mol); PPi → 2Pi drives irreversible reactions
• Colligative properties: depend on number of solute particles, not identity; osmotic pressure (πV = nRT); freezing point depression; boiling point elevation; relevant to osmosis across cell membranes

1.4 Bioenergetics and Metabolism Overview

• Glycolysis (Embden-Meyerhof-Parnas): glucose → 2 pyruvate; net yield 2 ATP + 2 NADH; key enzymes: Hexokinase (regulated by G-6-P), PFK-1 (rate-limiting; activated by AMP, F-2,6-BP; inhibited by ATP, citrate), Pyruvate kinase
• TCA cycle (Krebs cycle): Acetyl-CoA + OAA → citrate → (8 steps) → OAA; per turn: 3 NADH + 1 FADH₂ + 1 GTP + 2 CO₂; irreversible steps: isocitrate DH (rate-limiting), α-KG DH, citrate synthase
• Oxidative phosphorylation: NADH → Complex I → CoQ → Complex III → Cyt c → Complex IV → O₂ (½ → H₂O); FADH₂ → Complex II → CoQ; Proton gradient drives ATP synthase (Complex V); P/O ratio: NADH = 2.5 ATP, FADH₂ = 1.5 ATP
• Total ATP from glucose: ~30–32 ATP (2 glycolysis + 2 TCA + ~26–28 oxidative phosphorylation)
• Fatty acid β-oxidation: activation (acyl-CoA); transport across inner mitochondrial membrane (carnitine shuttle); spiral of 4 reactions (oxidation-hydration-oxidation-thiolysis); each 2C removed → 1 FADH₂ + 1 NADH + 1 acetyl-CoA; palmitate (16C) → 8 acetyl-CoA + 7 FADH₂ + 7 NADH → ~106 ATP
• Pentose phosphate pathway (PPP): G-6-P → ribose-5-P (for nucleotide synthesis) + NADPH (for reductive biosynthesis, antioxidant — regenerates glutathione); G6PD deficiency → haemolytic anaemia with oxidant drugs/fava beans

1.5 Enzymes and Enzyme Kinetics

• Enzymes: biological catalysts; lower activation energy (Ea); not consumed; highly specific; regulated
• Active site: small region that binds substrate and catalyses reaction; amino acid residues determine specificity (Lock and Key model — rigid; Induced Fit model — flexible, conformational change upon substrate binding — more accurate)
• Michaelis-Menten kinetics: v = Vmax[S]/(Km + [S]); Km = [S] at ½Vmax (affinity measure — lower Km = higher affinity); Vmax = maximum velocity; kcat = turnover number (reactions/enzyme/sec); kcat/Km = catalytic efficiency
• Lineweaver-Burk (double reciprocal plot): 1/v vs 1/[S]; y-intercept = 1/Vmax; x-intercept = –1/Km; used to determine inhibitor type

• Allosteric regulation: effector binds distal allosteric site → conformational change → alters activity; cooperative binding (sigmoidal kinetics — Hill equation); homotropic (substrate itself) vs heterotropic (different molecule) allosterism
• Covalent modification: phosphorylation (Ser, Thr, Tyr — by kinases; reversed by phosphatases); acetylation; ubiquitination; glycosylation; ADP-ribosylation
• Zymogen activation: inactive precursor activated by proteolytic cleavage; examples: trypsinogen → trypsin (by enterokinase); pepsinogen → pepsin (acid pH); blood clotting cascade; caspases (apoptosis)
• Enzyme classes (EC numbering): 1 Oxidoreductases, 2 Transferases, 3 Hydrolases, 4 Lyases, 5 Isomerases, 6 Ligases (synthetases)
• Ribozymes: catalytic RNA; 23S rRNA (peptidyl transferase activity in ribosome); self-splicing introns (Group I and II); RNase P (tRNA processing)

2.1 Membrane Structure and Function

• Fluid Mosaic Model (Singer and Nicolson, 1972): lipid bilayer is fluid (lateral movement of lipids, "flip-flop" very rare); proteins embedded (integral) or attached (peripheral); cholesterol regulates fluidity
• Membrane lipid composition: phospholipids (majority); sphingolipids; cholesterol; glycolipids (outer leaflet only — sugar facing out); asymmetric distribution between leaflets
• Membrane proteins: Integral (transmembrane — single pass or multi-pass; glycophorin, ion channels, receptors, transporters); Peripheral (attached to surface by electrostatic/H-bonds; cytochrome c, spectrin)
• Membrane transport: Simple diffusion (small non-polar molecules — O₂, CO₂, N₂, steroid hormones; small uncharged polar — H₂O, glycerol); Facilitated diffusion (requires transporter or channel; glucose transporter GLUT, aquaporins, ion channels — no energy); Active transport (requires ATP; Primary — directly uses ATP, e.g., Na⁺/K⁺-ATPase; Secondary — uses ion gradient, e.g., Na⁺-glucose symport in intestine)
• Na⁺/K⁺-ATPase: 3 Na⁺ out, 2 K⁺ in per ATP; maintains resting membrane potential; inhibited by ouabain (cardiac glycoside)
• Osmosis: water moves across semipermeable membrane from low to high solute concentration (hypotonic → hypertonic); turgor pressure in plants; cell lysis in hypo-osmotic environment; crenation (animal cells) in hyper-osmotic environment

2.2 Organelles — Structure and Function

Nucleus

• Bounded by nuclear envelope (double membrane); nuclear pore complexes (NPCs) — 120 nm diameter; RanGTP gradient controls import/export
• Nucleoplasm contains: chromatin (euchromatin — transcriptionally active; heterochromatin — condensed, silent); nucleolus (rRNA synthesis and ribosome assembly; 45S pre-rRNA → 28S + 18S + 5.8S); nuclear lamina (Lamin A, B, C — provides structural support; mutations → Hutchinson-Gilford Progeria)

Mitochondria

• Double membrane; cristae (inner membrane folds — increase surface area for ATP synthesis); matrix (TCA cycle enzymes, mtDNA, ribosomes)
• Outer membrane: porin channels (VDAC) — large molecule passage; Inner membrane: impermeable (maintains proton gradient); electron transport chain complexes (I–IV) + ATP synthase (Complex V)
• ATP synthase (F₀F₁-ATPase): F₀ embedded in inner membrane (proton channel); F₁ protrudes into matrix (catalytic head); proton flow through F₀ rotates γ-subunit → drives ATP synthesis
• Mitochondrial DNA (mtDNA): 16.5 kb circular; 37 genes (13 polypeptides of ETC/ATP synthase, 22 tRNAs, 2 rRNAs); maternal inheritance; D-loop region (control region for replication/transcription); mtDNA mutations → MERRF, MELAS, Leber's hereditary optic neuropathy
• Endosymbiotic origin: α-proteobacterial ancestor; supported by double membrane, circular DNA, 70S ribosomes, binary fission

Chloroplast

• Triple membrane (outer, inner, thylakoid); stroma (dark reactions — Calvin cycle); thylakoid membrane (light reactions — PS I and PS II, ETC, ATP synthase); grana (stacked thylakoids); stroma lamellae (unstacked, connecting grana)
• Photosystems: PS II (P680 — chlorophyll absorbs at 680 nm; water splitting — O₂ evolution; plastoquinone accepts electrons); PS I (P700; NADP⁺ reduction; final electron acceptor is NADPH)
• Z-scheme: H₂O → PS II → PQ → Cyt b₆f → PC → PS I → Fd → FNR → NADPH; ATP made by photophosphorylation (proton gradient from stroma→thylakoid lumen)
• Calvin cycle (dark/light-independent reactions): CO₂ + RuBP → 2 PGA (via RuBisCo); PGA → G3P (uses ATP + NADPH from light reactions); G3P → RuBP (regeneration — uses ATP); net: 3 CO₂ → 1 G3P (requires 9 ATP + 6 NADPH)
• Photorespiration: RuBisCo adds O₂ to RuBP → phosphoglycolate (wasteful); C4 plants (maize, sugarcane) suppress by CO₂ concentration in bundle sheath cells (Hatch-Slack pathway); CAM plants (cacti) fix CO₂ at night

Endoplasmic Reticulum and Golgi

• Rough ER (RER): studded with ribosomes; site of synthesis of secretory/membrane proteins; N-glycosylation begins; protein folding (Hsp70/BiP, calnexin, calreticulin); quality control → misfolded proteins degraded (ERAD)
• Smooth ER (SER): no ribosomes; lipid synthesis (phospholipids, cholesterol, steroid hormones); drug detoxification (CYP450 enzymes in liver SER); Ca²⁺ store (IP₃-mediated Ca²⁺ release)
• Golgi apparatus: cis (receives from ER) → medial → trans (sorts and ships); O-glycosylation; glycolipid synthesis; protein sorting — default pathway is secretion; lysosomal proteins tagged with mannose-6-phosphate (M6P); vesicular trafficking: COP-II (ER→Golgi anterograde), COP-I (Golgi→ER retrograde), Clathrin (trans-Golgi→endosome→lysosome; receptor-mediated endocytosis)
• Lysosomes: acid hydrolases (active at pH 5); formed by fusion of late endosomes + vesicles from Golgi; phagocytosis (bacteria, dead cells), autophagy (damaged organelles); lysosomal storage diseases: Tay-Sachs (β-hexosaminidase A deficiency → GM₂ ganglioside accumulation), Gaucher (glucocerebrosidase), Niemann-Pick (sphingomyelinase)

Cytoskeleton

2.3 Cell Division — Mitosis and Meiosis

Mitosis

• Purpose: growth, repair, asexual reproduction; produces 2 genetically identical diploid daughter cells
• Prophase: chromatin condenses; nuclear envelope breaks down; spindle forms (from centrosomes in animals)
• Prometaphase: nuclear envelope gone; kinetochore microtubules attach to kinetochores; chromosomes begin moving
• Metaphase: chromosomes align at metaphase plate (plate is equidistant from poles); spindle assembly checkpoint (SAC) — ensures all kinetochores attached; Mad1, Mad2, BubR1 — delay anaphase until all chromosomes bi-oriented
• Anaphase: cohesins cleaved by separase (activated when SAC satisfied — securin degraded → separase active); sister chromatids pulled to poles; kinetochore microtubule shortening; polar microtubule elongation
• Telophase + Cytokinesis: nuclear envelopes reform; chromosomes decondense; cleavage furrow (animals — contractile ring of actin + myosin); cell plate (plants — vesicles fuse at equatorial plate)

Meiosis

• Purpose: sexual reproduction; produces 4 genetically distinct haploid daughter cells; two divisions (Meiosis I and II)
• Meiosis I (Reductional division): homologous chromosomes separate
• Prophase I substages: Leptotene (chromosomes condense), Zygotene (synapsis — homologs pair via synaptonemal complex), Pachytene (crossing over — chiasmata form; key stage for recombination), Diplotene (synaptonemal complex dissolves; chiasmata visible; lampbrush chromosomes in oocytes), Diakinesis (maximal condensation; terminalization of chiasmata)
• Metaphase I: bivalents (tetrads) align at plate; random assortment
• Anaphase I: homologs separate (cohesins cleaved except at centromere region — protected by Shugoshin/PP2A); sister chromatids remain joined
• Meiosis II: similar to mitosis; separates sister chromatids
• Significance of meiosis: Genetic diversity via crossing over (recombination) and independent assortment; halving of chromosome number (restored at fertilisation)

Cell Cycle and its Regulation

• Phases: G₁ (growth, preparation) → S (DNA synthesis) → G₂ (growth, check) → M (mitosis) → (G₀ — quiescence)
• Cyclins and CDKs: Cyclin D/CDK4,6 (G₁ — phosphorylate Rb); Cyclin E/CDK2 (G₁/S transition); Cyclin A/CDK2 (S phase); Cyclin A,B/CDK1 (G₂/M — MPF: M-phase Promoting Factor)
• Rb protein: tumour suppressor; unphosphorylated Rb binds E2F (transcription factor) → inhibits S-phase genes; CDK4/6-Cyclin D phosphorylates Rb → E2F released → transcribes genes for S phase entry
• Checkpoints: G₁/S checkpoint (DNA damage? → p53 → p21 → CDK inhibition → arrest); G₂/M checkpoint (DNA replicated?); Spindle Assembly Checkpoint (all chromosomes attached?)
• p53 — guardian of the genome: tumour suppressor; activated by DNA damage; induces p21 (CDK inhibitor → G₁ arrest), promotes DNA repair, induces apoptosis if damage irreparable; mutated in ~50% of all cancers

2.4 Chromosome Structure and Types

• Nucleosome: 147 bp DNA wound 1.7 turns around histone octamer (H2A, H2B, H3, H4 — two each); linked by 10–80 bp linker DNA + H1; "beads on a string" (10 nm fibre) → 30 nm fibre → 300 nm loops → 700 nm → 1400 nm (metaphase chromosome)
• Histone modifications: Acetylation (by HAT; removes positive charge; loosens DNA-histone → active transcription); Deacetylation (by HDAC → repression); Methylation (H3K4me3 — active; H3K9me3 — repression; H3K27me3 — Polycomb repression); Phosphorylation (H3S10-P — mitosis); Ubiquitination (H2A — repression; H2B — transcription)
• Polytene chromosomes: endoreduplication (DNA replication without cell division); homologs paired; 1000+ copies; puffs = actively transcribed regions; seen in Drosophila salivary glands; used for chromosome mapping
• Lampbrush chromosomes: diplotene stage of meiosis I in oocytes (amphibians especially); lateral loops extend from chromomeres; actively transcribing; can be visualised under light microscope
• B chromosomes: supernumerary chromosomes; not essential; found in many species; may confer some reproductive advantage

3.1 DNA Replication

• Semiconservative: each daughter DNA has one old and one new strand (Meselson-Stahl experiment 1958 — ¹⁵N/¹⁴N density gradient centrifugation)
• Origin of replication (ori): prokaryotes — single oriC; eukaryotes — multiple origins (ARS in yeast); initiator proteins bind ori → recruit helicase
• Eukaryotic replication machinery:

• Okazaki fragments: lagging strand synthesised discontinuously in 100–200 bp (eukaryotes) or 1000–2000 bp (prokaryotes) fragments; each needs new primer
• Telomere replication: conventional DNA pol cannot replicate 5' ends (end-replication problem → chromosome shortening); Telomerase (a ribonucleoprotein — telomerase RNA component TERC + TERT reverse transcriptase) extends 3' end; active in germ cells, stem cells, cancer cells; absent/inactive in most somatic cells → replicative senescence

DNA Damage and Repair

3.2 Transcription

• RNA synthesis: 5'→3' direction; template (antisense) strand read 3'→5'; no primer needed; uses NTPs (ribo-); one-strand of DNA used as template
• Prokaryotic transcription: single RNA pol (core: α₂ββ'ω; holoenzyme with σ factor); σ recognises –10 and –35 promoter elements (consensus: TATAAT at –10, TTGACA at –35); different σ factors for different conditions (σ⁷⁰ — housekeeping, σ³² — heat shock); termination: Rho-independent (hairpin loop + U-run) or Rho-dependent (ATP-dependent RNA helicase Rho)
• Eukaryotic transcription: three RNA polymerases (Pol I — rRNA 45S; Pol II — mRNA + most snRNA + miRNA; Pol III — tRNA, 5S rRNA, snRNA); Pol II requires general transcription factors (TFIID-TATA box binding, TFIIA, TFIIB, TFIIF, TFIIE, TFIIH — helicase + CTD kinase); Mediator complex connects activators to Pol II

mRNA Processing (Eukaryotes)

• 5' capping: 7-methylguanosine cap added co-transcriptionally; protects from exonuclease degradation; required for ribosome recognition and translation initiation
• 3' polyadenylation: cleavage after AAUAAA polyadenylation signal; poly(A) polymerase adds 200–250 A residues; stability; mRNA export
• Splicing: removal of introns; spliceosome (snRNPs: U1, U2, U4, U5, U6 snRNAs + proteins); two transesterification steps; branch point A in intron; lariat intermediate; alternative splicing → multiple protein isoforms from one gene (human: ~95% of multi-exon genes undergo AS)
• Alternative splicing types: Exon skipping (most common in vertebrates); Alternative 5' splice site; Alternative 3' splice site; Intron retention; Mutually exclusive exons
• RNA editing: A-to-I editing by ADAR (adenosine deaminase acting on RNA); C-to-U editing by APOBEC; changes codon identity; ApoB-100 → ApoB-48 by C-to-U editing in intestine

Gene Regulation

• Prokaryotic gene regulation: Operon model (Jacob and Monod, 1961); lac operon (inducible — glucose absent + lactose present → CAP-cAMP activates, repressor off); trp operon (repressible — tryptophan present → activates repressor → turns off); attenuation (trp operon — ribosome stalling at leader peptide)
• Eukaryotic cis-regulatory elements: Promoter (core: TATA box ~–25; Inr; DPE); Enhancers (can be far, looping); Silencers; Insulators (CTCF — boundary elements)
• Transcription factors: DNA-binding domains (Zinc finger, Leucine zipper/bZIP, Helix-turn-helix, Helix-loop-helix); activation domains (acidic, proline-rich, glutamine-rich)
• Chromatin remodelling: SWI/SNF complex (uses ATP to reposition nucleosomes); histone modifying enzymes (HATs, HDACs, HMTs, HDMs) → change accessibility to transcription machinery
• Epigenetic regulation: DNA methylation (5-methylcytosine at CpG islands — gene silencing; maintained by DNMT1 after replication; de novo methylation by DNMT3A/3B); histone modifications; non-coding RNAs
• RNA interference (RNAi): miRNA (encoded by genome; ~22 nt; processed by Drosha/DGCR8 in nucleus → Dicer in cytoplasm → RISC complex; partial complementarity → mRNA degradation or translational repression); siRNA (exogenous dsRNA; perfect complementarity → mRNA cleavage by Argonaute); piRNA (germ cell transposon silencing)

3.3 Translation

• Genetic code: 64 codons for 20 amino acids + 3 stop codons (UAA, UAG, UGA); degenerate/redundant (multiple codons per AA; wobble at 3rd position — Crick 1966); universal (few exceptions — mitochondria, Mycoplasma); non-overlapping; unambiguous (each codon = one AA)
• Start codon: AUG (Met in eukaryotes; fMet in prokaryotes); Kozak sequence context in eukaryotes (GCCRCCAUGG, R = purine)
• tRNA: adaptor molecule; anticodon on anticodon loop; amino acid attached to CCA-3' by aminoacyl-tRNA synthetase (20 enzymes — one per AA; uses ATP; proofreading activity); wobble — non-standard base pairing at 3rd codon position (inosine most flexible)

Translation Steps

• Polysome (polyribosome): multiple ribosomes translating same mRNA simultaneously — increases efficiency
• Protein targeting: Signal Hypothesis (Blobel and Sabatini); signal peptide (N-terminal hydrophobic); SRP (Signal Recognition Particle) recognises signal → docks with SRP receptor on RER → co-translational insertion into ER
• Post-translational modifications: signal peptide cleavage; glycosylation; phosphorylation; acetylation; ubiquitination (26S proteasome degradation); prenylation; GPI anchoring; disulfide bond formation (in ER lumen — PDI catalyses)
• Protein folding and chaperones: Hsp70 (BiP in ER) — binds hydrophobic segments of unfolded proteins; GroEL/GroES (Hsp60) — barrel-shaped cage for folding; Hsp90 — client proteins include steroid hormone receptors, kinases
• UPR (Unfolded Protein Response): triggered by ER stress (accumulation of misfolded proteins); sensors: IRE1, PERK, ATF6 → reduce translation + increase chaperone production; if unresolved → apoptosis

3.4 DNA Topology and the Modern Concept of the Gene

• DNA topology: supercoiling (over/under-winding); negative supercoiling (underwound — in bacteria and eukaryotic chromatin) facilitates strand separation for replication and transcription; positive supercoiling ahead of replication fork; topoisomerases resolve
• DNA motifs: TATA box, CAAT box, GC box, E-box, CpG islands (often near promoters — hypomethylated in active genes)
• Modern gene concept: gene = DNA sequence specifying functional product (RNA or protein); includes regulatory sequences, introns, UTRs; one gene can produce multiple products via alternative splicing, alternative promoters, alternative polyadenylation, RNA editing
• Split genes (interrupted genes): discovered by Roberts and Sharp (1977 Nobel 1993); introns interrupt coding sequences; mosaic genes; exon shuffling in evolution
• Pseudogenes: non-functional gene copies; processed pseudogenes (retrotransposed cDNA copies); duplicated pseudogenes (lost function by mutation)
• Non-coding RNA genes: miRNA, siRNA, lncRNA, piRNA, circRNA — transcribed but not translated; regulatory functions; much of "junk DNA" is transcribed into ncRNA

4.1 Principles of Cell Signaling

• Types of signaling: Endocrine (hormones via blood — long range); Paracrine (local diffusion — cytokines, growth factors); Autocrine (cell responds to own signals — common in cancer); Juxtacrine (contact-dependent — Notch-Delta, EGF ligand)
• Signal transduction steps: Ligand binding → receptor activation → transduction (second messengers, kinase cascades) → effector response → signal termination
• Second messengers: cAMP (adenylyl cyclase → PDE degrades); cGMP; IP₃ (releases Ca²⁺ from ER); DAG (activates PKC); Ca²⁺ (binds calmodulin → CaM kinase II, MLCK); nitric oxide (NO — activates soluble guanylyl cyclase)

Receptor Classes

4.2 Key Signaling Pathways

Ras/MAPK Pathway

• RTK activation → Grb2-SOS complex recruited → SOS (GEF) activates Ras (GDP→GTP) → Ras-GTP → Raf (MAPKKK) → MEK (MAPKK) → ERK (MAPK) → transcription factors (Elk-1, Fos, c-Myc) → cell proliferation
• Ras is a GTPase — intrinsic GTPase activity terminates signaling (GAP proteins accelerate GTP hydrolysis); Ras mutations (G12V or Q61L) — oncogenic (constitutively active); mutated in ~30% cancers (K-Ras most common)

PI3K/Akt/mTOR Pathway

• RTK → PI3K phosphorylates PIP₂ → PIP₃ → recruits PDK1 and Akt (PKB) → Akt phosphorylates: mTOR (protein synthesis, cell growth), FOXO (survival), BAD (anti-apoptosis), GSK3β (glycogen synthesis), MDM2 (p53 degradation)
• PTEN (tumour suppressor): lipid phosphatase → dephosphorylates PIP₃ back to PIP₂ → negative regulator; mutated in many cancers (Cowden syndrome)
• mTOR: master growth regulator; mTORC1 (rapamycin-sensitive) → phosphorylates S6K and 4E-BP1 → protein synthesis, ribosome biogenesis, autophagy inhibition

Wnt/β-catenin Pathway

• Without Wnt: destruction complex (APC + Axin + GSK3β + CK1) phosphorylates β-catenin → ubiquitination → proteasomal degradation; TCF/LEF represses Wnt target genes
• With Wnt: Frizzled + LRP5/6 → Dishevelled → inhibits destruction complex → β-catenin accumulates → translocates to nucleus → TCF/LEF-β-catenin activates target genes (c-Myc, Cyclin D1, MMP-7) → cell proliferation
• APC mutation: most common in colorectal cancer (FAP — Familial Adenomatous Polyposis); loss of APC → uncontrolled β-catenin → unregulated proliferation

Notch Signaling

• Juxtacrine signaling; Notch receptor (transmembrane) on receiving cell; Delta/Jagged ligand on signaling cell
• Ligand binding → γ-secretase cleavage → Notch intracellular domain (NICD) released → enters nucleus → CSL transcription factor complex → Hes/Hey target genes → cell fate decisions (binary — neurogenesis, somitogenesis, lateral inhibition)

Signal Termination and Desensitisation

• Receptor phosphorylation by GRK (GPCR Kinase) → β-arrestin recruitment → receptor internalisation (clathrin-mediated) → receptor recycling or degradation
• Phosphodiesterase (PDE): degrades cAMP and cGMP; inhibited by caffeine (mild PDE inhibitor — explains mood effects)
• Phosphatases: protein phosphatases (PP1, PP2A, PP2B/calcineurin — target of cyclosporin in immunosuppression; PP2C); tyrosine phosphatases (PTP1B — insulin resistance target)

5.1 Embryogenesis and Early Development

• Fertilisation: sperm binds ZP3 on zona pellucida → acrosome reaction → penetration → sperm-egg membrane fusion → fast block to polyspermy (membrane depolarisation); slow block (cortical reaction — cortical granules fuse with egg membrane → zona reaction → permanent block); egg activation → meiosis II completion
• Cleavage: rapid mitotic divisions with no growth; maintains cytoplasm:nucleus ratio; holoblastic (entire egg cleaves — sea urchin, frog, mammal); meroblastic (partial — yolk-rich eggs; discoidal in fish, rotational in birds)
• Morula → Blastocyst (mammal): ICM (inner cell mass — gives embryo) + trophectoderm (gives placenta); implantation; OCT4, SOX2, NANOG — pluripotency factors in ICM

Establishment of Body Axes

• Drosophila body axes: A-P axis — Bicoid mRNA (anterior pole → Bicoid protein gradient → activates anterior gap genes like hunchback; inhibits posterior); Nanos mRNA (posterior → represses bicoid-type translation); D-V axis — Toll receptor gradient; terminal — Torso RTK
• Vertebrate A-P axis: Organiser (Spemann's organiser in amphibians — transplant → induces secondary axis; Hensen's node in chick; node in mouse); secretes inhibitors of BMP (Chordin, Noggin, Follistatin) → dorsal fate; BMP gradient → ventral fate; D-V axis established by BMP gradient
• Homeotic genes (Hox genes): define segment identity along A-P axis; discovered in Drosophila (homeotic mutations — antennapedia, bithorax); co-linearity — Hox genes expressed in same order along chromosome as their A-P expression domain; extremely conserved from Drosophila to humans; 39 Hox genes in humans (4 clusters: HOXA-D)
• Morphogens: signaling molecules that form concentration gradients → different cell fates at different concentrations; Bicoid, Activin/Nodal, Sonic Hedgehog (SHH), BMP, Wnt, Retinoic acid

Gastrulation

• Blastula → gastrula; establishment of three germ layers
• Frog gastrulation: dorsal blastopore lip (future notochord/Spemann organiser) involutes; bottle cells; three germ layers form
• Chick gastrulation: epiblast cells migrate through primitive streak → ingress; hypoblast → endoderm; epiblast → ectoderm and mesoderm after ingression; Hensen's node at anterior end of primitive streak = chick organiser

Fate Map and Cell Lineage

• Fate map: mapping regions of blastula/gastrula to their adult derivatives; using dye injection or lineage tracing (Cre-lox); Vogt developed vital dye technique
• Germ layers and derivatives: Ectoderm → epidermis, nervous system (neural induction — notochord secretes signals), sense organs, neural crest; Mesoderm → muscles, bones, blood, kidneys, gonads, heart; Endoderm → gut, liver, pancreas, lungs, thyroid
• Neural crest: "4th germ layer"; migratory cells from dorsal neural tube; forms: peripheral nervous system, craniofacial cartilage/bone, melanocytes, adrenal medulla (chromaffin cells), cardiac outflow tract; EMT (epithelial-to-mesenchymal transition) allows migration

5.2 Organogenesis — Eye and Heart Development

Eye Development

• Optic vesicle (from diencephalon) induces overlying ectoderm → lens placode → lens vesicle; optic vesicle invaginates → optic cup (bilayered: inner = neural retina, outer = RPE)
• Pax6 (Master control gene for eye): loss → no eye (eyeless in Drosophila); Pax6 overexpression anywhere → ectopic eye; highly conserved; induces entire eye developmental programme
• Retina specification: RGCs (retinal ganglion cells — only neurons that project axons to brain); rods (scotopic — dim light, rhodopsin); cones (colour, bright light — L/M/S cones); Müller glia; bipolar cells; amacrine; horizontal

Heart Development

• First organ to function in embryo; cardiomyocytes from lateral plate mesoderm (cardiac progenitor cells — FHF and SHF); heart tube forms and loops (rightward — D-looping); cardiac jelly; cardiac neural crest (great vessel formation)
• Key TFs: Nkx2.5 (homologue of tinman in Drosophila — required for heart formation), GATA4, Hand1/2, MEF2C, T-box factors (Tbx5 — mutations → Holt-Oram syndrome; Tbx20)
• Common congenital heart defects: VSD (ventricular septal defect — most common), ASD, tetralogy of Fallot, transposition of great arteries, patent ductus arteriosus

5.3 Developmental Genetics and Regeneration

• Genetic vs induced teratogenesis: Genetic — chromosomal anomalies (Down, Turner, Klinefelter), single gene mutations; Induced — teratogens (thalidomide — phocomelia via Wnt/CRBN; valproate — NTDs; isotretinoin — retinoic acid excess → craniofacial); radiation; infection (rubella → cardiac defects, deafness, cataracts)
• Thyroxine and metamorphosis in amphibia: thyroid hormone (T₃/T₄) drives metamorphosis in frogs; tail resorption (apoptosis), limb growth, lung development, gut remodelling; each tissue responds differently to same hormone (different receptor isoforms, local factors); hypothyroidism → arrested metamorphosis (neoteny in axolotl)
• Paedogenesis: sexual reproduction in larval stage (e.g., some insects); Neoteny: retention of juvenile features in adult; axolotl (Ambystoma mexicanum) — neotenic salamander; fails to metamorphose due to low thyroid sensitivity; can be induced by T₄
• Regeneration: replace lost body parts; Epimorphosis (de-differentiation then re-differentiation — Planaria, newt limb); Morphallaxis (remodelling without cell division — Hydra); Compensatory growth (liver regeneration — remaining hepatocytes divide); Stem cell-mediated (replacement from resident stem cells)
• Newt lens regeneration: only vertebrate that can regenerate lens; from pigmented epithelial cells of dorsal iris (transdifferentiation — mature differentiated cells convert to another type); Wnt, FGF, retinoic acid pathways
• Cell death (apoptosis in development): Apoptosis sculpts morphology; interdigital spaces (webbing removal); matching neuron numbers to targets (neurotrophic hypothesis — NGF withdrawal → apoptosis); thymus T cell selection; deletion of autoreactive lymphocytes
• Ageing: Telomere shortening; accumulation of ROS; mitochondrial dysfunction; senescent cells (SASP — senescence-associated secretory phenotype — chronic inflammation); epigenetic drift; NAD⁺ depletion; stem cell exhaustion; proteostasis failure

In Vitro Fertilisation and Embryo Transfer

• IVF: ovarian stimulation → oocyte retrieval → fertilisation in vitro → embryo culture → transfer to uterus; India: first test tube baby Durga (1978 Calcutta) and UK Louise Brown (1978, Robert Edwards — Nobel 2010)
• ICSI (Intracytoplasmic Sperm Injection): single sperm injected into egg; for severe male infertility
• PGD (Preimplantation Genetic Diagnosis): biopsy of 1–2 blastomeres from Day 3 embryo (8-cell); genetic testing before transfer; screens for chromosomal/single gene disorders
• Embryo cryopreservation: vitrification (rapid freezing) used now; long-term storage; donor egg programmes

Cloning and Stem Cells

• Somatic Cell Nuclear Transfer (SCNT): nucleus from somatic cell → enucleated oocyte → reprogramming by oocyte cytoplasm → cloned embryo; Dolly the sheep (Wilmut, 1996 — first cloned mammal from adult somatic cell); very inefficient (~1% success rate)
• iPSCs (Induced Pluripotent Stem Cells): Yamanaka (2006 Nobel 2012 with Gurdon); fibroblasts + 4 factors (Oct4, Sox2, Klf4, c-Myc or Oct4, Sox2, Lin28, Nanog) → reprogrammed to pluripotent stem cells; avoids ethical issues of ESC
• Stem cell types: Totipotent (zygote — can form all embryonic and extraembryonic tissues); Pluripotent (ICM, ESC, iPSC — all 3 germ layers but not placenta); Multipotent (HSC, MSC, NSC — limited lineages); Unipotent (satellite cells in muscle)
• Therapeutic uses of stem cells: Bone marrow transplant (HSC); skin grafting (epidermal stem cells); corneal transplant; clinical trials for Parkinson's, diabetes (β-cell replacement), spinal cord injury, ALS

6.1 Photosynthesis — Light and Dark Reactions

• Photosynthesis: 6CO₂ + 6H₂O + light → C₆H₁₂O₆ + 6O₂; two stages: light-dependent (thylakoid) + light-independent/Calvin cycle (stroma)
• Light reactions: Photon absorption by antenna pigments → funnelled to reaction centre chlorophyll; PS II (P680) — excited by light → oxidises water (Mn cluster — OEC, oxygen evolving complex → O₂); electrons → PQ → Cyt b₆f (pumps H⁺ → lumen) → PC → PS I (P700) → ferredoxin (Fd) → FNR → NADPH
• Cyclic photophosphorylation: PS I → Fd → Cyt b₆f → PC → PS I; produces only ATP (no NADPH, no O₂); used when NADPH/ATP ratio is high
• Photophosphorylation: ATP synthesis driven by proton gradient (lumen to stroma) through ATP synthase (CF₀CF₁); ~3 H⁺ per ATP
• Calvin cycle (C3 pathway): CO₂ + RuBP → 2 PGA (3C) via RuBisCO; PGA → G3P (3 ATP + 2 NADPH per CO₂ fixed for 6 turns to make 1 G3P net); regeneration of RuBP (9 ATP for 3 CO₂ fixed)

C4 and CAM Plants

6.2 Plant Hormones

6.3 Plant Nutrition and Transport

Mineral Nutrition

• Essential mineral elements: macro (N, P, K, Ca, Mg, S — required in large amounts); micro/trace (Fe, Mn, Zn, Cu, B, Mo, Cl, Ni — required in small amounts)
• Nitrogen fixation: biological (by Rhizobium symbiosis in root nodules of legumes; free-living Azotobacter, Anabaena, Cyanobacteria); industrial (Haber-Bosch — N₂ + 3H₂ → 2NH₃); Nitrogenase enzyme (Fe-Mo protein + Fe protein; ATP-dependent; sensitive to O₂ → leghemoglobin in nodules protects nitrogenase)
• Nitrogen cycle: N₂ fixation → NH₄⁺ (ammoniation) → NO₂⁻ → NO₃⁻ (nitrification by Nitrosomonas, Nitrobacter); NO₃⁻ → N₂ (denitrification — anaerobic bacteria); organic N → NH₄⁺ (ammonification/mineralisation)
• Mineral deficiency symptoms: N (chlorosis of older leaves), P (purple coloration — anthocyanin), K (leaf margin scorch), Ca (poor root growth; blossom end rot), Fe (chlorosis of young leaves — not re-mobilised), Mg (interveinal chlorosis — mobile), Mn (grey speck in oats), Zn (little leaf, rosette), B (death of growing points)

Water and Solute Transport

• Water potential (Ψw) = Ψs (solute/osmotic potential — always negative) + Ψp (pressure potential); water moves from high Ψw to low Ψw
• Pathways in root: Apoplastic (through cell walls — blocked at Casparian strip of endodermis for ions); Symplastic (through cytoplasm and plasmodesmata); Transmembrane (through cell membranes + plasmodesmata)
• Casparian strip: suberin-impregnated strip in endodermal cell walls; forces all water and ions through symplast → selective uptake control
• Transpiration pull: evaporation from mesophyll cells → negative pressure in xylem → water pulled up (SPAC — Soil-Plant-Atmosphere Continuum); Cohesion-Tension theory (Dixon and Joly 1894)
• Stomatal aperture control: K⁺ uptake into guard cells → water uptake → guard cell swelling → stomata open; ABA reverses by causing K⁺ efflux; CO₂ sensors; blue light (phototropin receptor) opens stomata; circadian rhythm
• Phloem transport (source → sink): sucrose loaded into phloem (companion cells → sieve elements) at source (mature leaf) → pressure-flow hypothesis — high turgor at source, low at sink → mass flow; unloading at sink (root, fruit, seed)

6.4 Plant Responses to Environment

• Photoperiodism: flowering response to relative length of light and dark periods; Long-Day Plants (LDP — flower when night < critical length: spinach, wheat); Short-Day Plants (SDP — flower when night > critical length: soybean, rice, chrysanthemum); Day-neutral plants (tomato, maize)
• Phytochrome (Pr/Pfr system): Pr (absorbs red light 660 nm → Pfr); Pfr (absorbs far-red 730 nm → Pr); Pfr promotes flowering in LDP, inhibits in SDP; night break effect; cytoplasmic and nuclear signaling (PIF — phytochrome interacting factors)
• Vernalisation: requirement for prolonged cold (0–7°C) to flower; epigenetic — FLC gene silenced by H3K27me3 during cold → FT expressed → flowering
• Photomorphogenesis: skotomorphogenesis (dark — etiolated, long hypocotyl, closed cotyledons); photomorphogenesis (light — short hypocotyl, open green cotyledons, COP1 ubiquitin ligase degraded in light)
• Tropisms: Phototropism (towards light — phototropin receptor in shoots; auxin redistribution); Gravitropism (roots: statoliths in columella cells fall to lower side → auxin redistribution → lower side grows slower in roots; shoots — opposite response); Thigmotropism (touch response — tendril coiling; seismonasty in Mimosa)
• Seed dormancy and germination: ABA enforces dormancy; GA breaks dormancy; light (phytochrome for small seeds); stratification (cold treatment); scarification; imbibition → activation of enzymes → α-amylase production (GA-dependent in aleurone)

7.1 Blood and Circulatory System

• Blood components: Plasma (55% — water, proteins: albumin [oncotic pressure], globulins [antibodies], fibrinogen [clotting], clotting factors, hormones, nutrients); RBCs/Erythrocytes (45% — biconcave disc, no nucleus, no mitochondria in mammals; haemoglobin; 120 day lifespan; destroyed in spleen); WBCs/Leucocytes; Platelets (thrombocytes — clotting)
• Haemoglobin (Hb): α₂β₂ tetramer; Fe²⁺-porphyrin (haem) in each subunit; cooperative O₂ binding (sigmoidal curve — Hill coefficient n=2.8); T state (deoxy — low O₂ affinity); R state (oxy — high O₂ affinity)
• Bohr effect: ↑ CO₂ or ↓ pH → right shift of O₂-Hb dissociation curve → ↓ O₂ affinity → O₂ released to respiring tissues; H⁺ + Hb-O₂ → Hb-H⁺ + O₂
• 2,3-BPG (bisphosphoglycerate): allosteric effector; binds β-chain cleft of deoxy-Hb (T state); right shifts curve; ↑ at high altitude → helps release O₂; fetal Hb (HbF, α₂γ₂) binds 2,3-BPG less → higher O₂ affinity than maternal HbA → O₂ transfer across placenta
• CO₂ transport: 70% as HCO₃⁻ (CO₂ + H₂O ⇌ H₂CO₃ ⇌ H⁺ + HCO₃⁻ — catalysed by carbonic anhydrase in RBCs; Cl⁻/HCO₃⁻ exchange — chloride shift); 23% as carbaminohaemoglobin; 7% dissolved in plasma
• Blood clotting cascade: Extrinsic (TF/Factor VII — activated by vessel injury) and Intrinsic (Factor XII/XI/IX/VIII — contact activation) pathways → Common pathway (X → Xa → prothrombin → thrombin → fibrinogen → fibrin → Factor XIII crosslinks fibrin); Vitamin K required for factors II, VII, IX, X (γ-carboxylation)
• Anticoagulants: Heparin (activates antithrombin III); Warfarin (Vitamin K antagonist); Aspirin (inhibits COX → reduced thromboxane A₂ → less platelet aggregation); tPA (tissue plasminogen activator — breaks clots → plasmin → fibrinolysis)

Cardiovascular Physiology

• Cardiac cycle: Systole (contraction) → diastole (relaxation); SA node (pacemaker — 70 bpm) → AV node (delay 0.1s) → Bundle of His → Purkinje fibres → ventricular muscle; action potential: depolarisation (Na⁺ fast influx) → plateau (Ca²⁺ influx from L-type channels → triggers SR Ca²⁺ release — CICR → contraction) → repolarisation (K⁺ efflux)
• Cardiac output: CO = Stroke Volume × Heart Rate; Starling's Law — increased venous return → increased stretch of ventricle → increased force of contraction (heterometric autoregulation)
• Blood pressure: systolic/diastolic; mean arterial pressure = diastolic + 1/3 pulse pressure; regulated by baroreceptors (aortic arch, carotid sinus) → autonomic nervous system + renin-angiotensin-aldosterone system (RAAS): renin (juxtaglomerular cells) → angiotensinogen → Ang I → ACE → Ang II → vasoconstriction + aldosterone (Na⁺/water retention) → ↑ BP

7.2 Digestive System

• Digestion: Mouth (salivary amylase — starch → dextrins; lysozyme); Stomach (HCl + pepsin — protein denaturation and pepsin activation; gastric lipase; intrinsic factor for B₁₂); Small intestine (main site); Large intestine (water absorption; microbial fermentation; vitamins K and B₁₂ from bacteria)
• Pancreatic enzymes (secreted as zymogens): Trypsinogen → trypsin (by enterokinase/enteropeptidase); trypsin activates all others: chymotrypsinogen → chymotrypsin; proelastase → elastase; procarboxypeptidase A/B → carboxypeptidase; pancreatic lipase (requires colipase); pancreatic amylase; ribonuclease, DNase
• Bile: made by liver, stored in gallbladder; bile salts (cholate, deoxycholate — emulsify fats → micelle formation → lipase access); bilirubin (haem catabolism product — conjugated by UDP-glucuronosyltransferase → water-soluble; excreted in bile → stercobilin in faeces; some re-absorbed → urobilinogen in urine)
• Fat absorption: long-chain FA + monoacylglycerol → chylomicron (assembled in intestinal cell) → lymph → blood; medium chain FA directly into portal blood; fat-soluble vitamins absorbed with fats
• GI hormones: Gastrin (G cells of stomach antrum — stimulates HCl, pepsin; stimulated by food, vagus); CCK (I cells of duodenum — gallbladder contraction, pancreatic enzyme secretion; stimulated by fat + protein); Secretin (S cells — pancreatic HCO₃⁻ secretion; stimulated by acid); GIP (K cells — insulin secretion; inhibits gastric motility); GLP-1 (L cells — incretin effect; GLP-1 receptor agonists like semaglutide for diabetes/obesity)
• Energy balance and BMR: Hypothalamus regulates appetite; Leptin (adipocyte — satiety); Ghrelin (stomach — hunger — rises before meals); Insulin (anabolic; promotes fat storage); NPY/AgRP (orexigenic hypothalamic neurons); POMC/CART (anorexigenic)

7.3 Respiratory System

• Lungs: right lung (3 lobes), left lung (2 lobes); bronchi → bronchioles → alveoli (300 million, surface area ~70 m²); type I pneumocytes (gas exchange); type II pneumocytes (surfactant — dipalmitoylphosphatidylcholine — reduces surface tension, prevents collapse)
• Respiratory mechanics: inspiration (active — diaphragm + external intercostals contract → ↑ thoracic volume → ↓ pressure → air in); expiration (passive at rest — elastic recoil; active in forced expiration — internal intercostals + abdominals)
• Lung volumes: Tidal Volume (TV) ~500 mL; Inspiratory Reserve Volume (IRV) ~3000 mL; Expiratory Reserve Volume (ERV) ~1200 mL; Residual Volume (RV) ~1200 mL (cannot be expelled); Vital Capacity (VC) = TV + IRV + ERV; Total Lung Capacity (TLC) = VC + RV
• Gas exchange: Fick's law: rate = (P₁–P₂) × A × D/T (diffusing capacity); partial pressure gradients: alveolar pO₂ ~100 mmHg → pulmonary capillary ~40 mmHg (venous) → equilibrates to ~100 mmHg; pCO₂ reverse gradient
• Control of breathing: Pre-Bötzinger complex (medullary rhythm generator); chemoreceptors: Central (medullary surface — responds to pH/pCO₂; pCO₂ is primary stimulus); Peripheral (carotid and aortic bodies — respond to pO₂, pCO₂, pH; hypoxic drive)
• Respiratory disorders: Obstructive (↓ FEV₁/FVC ratio — asthma, COPD, emphysema); Restrictive (↓ FVC, normal FEV₁/FVC — pulmonary fibrosis, sarcoidosis)

7.4 Excretory System

• Kidney anatomy: cortex (glomeruli, PCT, DCT); medulla (loop of Henle, collecting duct); nephron (functional unit — ~1 million per kidney)
• Nephron function — four processes:

• Acid-base balance: kidneys excrete H⁺ (as NH₄⁺ and titratable acid) and reabsorb HCO₃⁻; Bicarbonate buffer is main blood buffer; respiratory compensation within minutes; renal compensation takes days
• Acid-base disorders: Metabolic acidosis (↓ pH, ↓ HCO₃⁻ — diabetic ketoacidosis, renal failure); Metabolic alkalosis (↑ pH, ↑ HCO₃⁻ — vomiting, diuretics); Respiratory acidosis (↓ pH, ↑ pCO₂ — COPD, hypoventilation); Respiratory alkalosis (↑ pH, ↓ pCO₂ — hyperventilation, high altitude)
• Juxtaglomerular apparatus: JG cells (modified smooth muscle of afferent arteriole — secrete renin); macula densa (DCT cells sensing Na⁺ delivery — regulate renin and GFR); mesangial cells; regulates GFR via tubuloglomerular feedback

7.5 Nervous System

• Neuron structure: cell body (soma) + dendrites (receive signals) + axon (transmit to presynaptic terminal); myelinated axons (Schwann cells in PNS; oligodendrocytes in CNS; saltatory conduction at nodes of Ranvier)
• Resting membrane potential: –70 mV (inside negative); maintained by Na⁺/K⁺-ATPase (3 Na⁺ out, 2 K⁺ in) + selective K⁺ permeability (K⁺ leaks out through K⁺ leak channels)
• Action potential: Threshold (–55 mV) → rapid depolarisation (voltage-gated Na⁺ channels open → Na⁺ influx) → overshoot (+40 mV) → repolarisation (Na⁺ channels inactivate; voltage-gated K⁺ channels open → K⁺ efflux) → undershoot/hyperpolarisation (K⁺ channels slow to close) → refractory period (absolute: Na⁺ channels inactivated; relative: hyperpolarised); All-or-none principle
• Synaptic transmission: Action potential → Ca²⁺ influx (VGCC) → vesicle fusion (SNARE proteins: SNAP-25, Synaptobrevin, Syntaxin) → neurotransmitter release → diffuses across cleft → postsynaptic receptor → response; neurotransmitter termination: reuptake (SERT for serotonin — SSRIs block; DAT for dopamine), enzymatic degradation (AChE for ACh), diffusion

Sense Organs

• Vision: Rods (rhodopsin — 11-cis retinal + opsin; absorbs ~500 nm; dim light/scotopic; no colour); Cones (three types: L/M/S — red/green/blue; photopic; colour vision); Phototransduction: light → 11-cis retinal → all-trans retinal → opsin activation → transducin (Gαt) → activates PDE → ↓ cGMP → cGMP-gated channels close → hyperpolarisation (unique — light causes hyperpolarisation not depolarisation) → reduced glutamate release → bipolar cells → ganglion cells → optic nerve
• Hearing: Sound waves → tympanic membrane → ossicles (malleus-incus-stapes) → oval window → cochlea (scala vestibuli-scala media-scala tympani); basilar membrane vibrates — tonotopic organisation (base = high freq; apex = low freq); hair cells (stereocilia) — tip links open mechanosensitive channels → K⁺ influx (endolymph is K⁺ rich) → depolarisation → glutamate release → auditory nerve (cranial nerve VIII)
• Tactile response: Meissner's corpuscles (light touch, edge detection); Pacinian corpuscles (vibration, deep pressure); Ruffini endings (skin stretch); Merkel discs (fine touch, sustained pressure); Free nerve endings (pain, temperature)

7.6 Endocrine System

7.7 Reproductive System and Physiology

• Menstrual cycle (28 days): Follicular phase (Day 1–14): FSH → follicle development → oestrogen ↑ → LH surge (mid-cycle, day 14) → Ovulation; Luteal phase (Day 14–28): LH → corpus luteum → progesterone + oestrogen ↑ → endometrium prepares; if no fertilisation → corpus luteum degenerates → progesterone ↓ → menstruation
• Puberty: GnRH pulse frequency increases → LH and FSH → gonadal steroid production; menarche (first menstruation); spermarche (first ejaculation); Tanner stages
• Menopause: ovarian follicle depletion → ↓ oestrogen and progesterone → ↑ FSH and LH (no negative feedback); hot flushes; osteoporosis risk; CVD risk increases; vaginal atrophy; average age 51 years
• Spermatogenesis: seminiferous tubules; spermatogonia → primary spermatocyte (meiosis I) → secondary spermatocyte (meiosis II) → spermatids → spermatozoa (spermiogenesis); ~70–74 days; temperature-sensitive (3°C below body — cryptorchidism → infertility)
• Oogenesis: all primary oocytes present at birth (arrested in Prophase I); resume meiosis at puberty (one per cycle); arrested at Metaphase II until fertilisation; asymmetric divisions → large egg + small polar bodies; only ~400 oocytes ovulated in lifetime
• Fertilisation and implantation: capacitation of sperm; acrosome reaction at zona pellucida; sperm-egg fusion; blocks to polyspermy; zygote → cleavage → morula → blastocyst → implants in endometrium (Day 6–7); hCG produced by trophoblast → maintains corpus luteum → early pregnancy
• Placenta: chorioallantoic; chorionic villi exchange O₂, nutrients, waste; barrier to large molecules, many pathogens (but rubella, CMV, Toxoplasma, HIV cross); produces oestrogen, progesterone (from 12 weeks → replaces corpus luteum), hCG, hPL

8.1 Mendelian Genetics

• Mendel's Laws: Law of Segregation (two alleles of a gene separate during gamete formation; each gamete receives one allele); Law of Independent Assortment (alleles of different genes assort independently — only true for non-linked genes)
• Monohybrid cross: Aa × Aa → 1 AA : 2 Aa : 1 aa (genotype 1:2:1; phenotype 3:1 for complete dominance)
• Test cross: Aa × aa → 1 Aa : 1 aa (1:1 ratio — identifies whether dominant phenotype is AA or Aa)
• Dihybrid cross: AaBb × AaBb → 9:3:3:1 (A_B_ : A_bb : aaB_ : aabb)

Extensions to Mendelian Genetics

8.2 Linkage, Recombination and Chromosome Mapping

• Linkage: genes on same chromosome tend to be inherited together (violate Law of Independent Assortment); coupling vs repulsion; linkage detected by deviation from 1:1:1:1 ratio in test cross of dihybrid
• Recombination frequency: RF = (recombinant progeny / total progeny) × 100%; 1% RF = 1 centimorgan (cM) = 1 map unit; RF maximum = 50% (for unlinked genes or genes far apart)
• Three-point cross: order three genes by determining which gene is in middle (double crossovers reveal middle gene); calculate map distances from each interval; positive interference — observed DCO less than expected; I = 1 – (observed DCO/expected DCO)
• Chromosome mapping: uses recombination frequencies between pairs of genes; maps are not always additive (double crossovers); LOD score (Log of Odds ratio) for human linkage analysis — threshold >3.0 suggests linkage

8.3 Sex Determination and Sex-Linked Inheritance

• XX/XY system (humans, Drosophila): SRY gene on Y chromosome → TDF (testis-determining factor) → male development; XO (Turner syndrome — female phenotype, sterile); XXY (Klinefelter — male phenotype, infertile); XXX (triple X — normal female)
• ZW/ZZ system (birds, some fish and insects): females are ZW; males are ZZ; opposite of XX/XY
• Haplo-diploid sex determination (Hymenoptera — bees, ants, wasps): males from unfertilised eggs (haploid); females from fertilised eggs (diploid); determines sex by ploidy, not sex chromosomes
• X-linked recessive inheritance: haemophilia A (Factor VIII — F8 gene); haemophilia B (Factor IX); colour blindness (OPN1LW, OPN1MW); Duchenne muscular dystrophy (dystrophin); G6PD deficiency; more common in males (XY — only one copy); females can be carriers (XX — heterozygous)
• X-linked dominant inheritance: Fragile X syndrome (FMR1 gene — CGG repeat expansion > 200 — leading cause of inherited intellectual disability); Rett syndrome (MECP2); vitamin D-resistant rickets
• Y-linked (Holandric) inheritance: father → all sons; Y chromosome (SRY, AZF — azoospermia factor, testis-specific genes)
• X-inactivation (Lyon hypothesis): one X randomly inactivated in each somatic cell of female → Barr body (condensed inactive X); epigenetically maintained by XIST RNA coating inactive X + H3K27me3; mosaicism in X-linked traits (calico cat coat colour)
• Dosage compensation: equalise gene expression between XX and XY; achieved by X-inactivation in mammals; hyperactivation of single X in Drosophila (MSL complex doubles transcription rate of X in males); chromosome-wide transcriptional silencing in C. elegans

8.4 Chromosomal Abnormalities and Human Genetics

Numerical Abnormalities (Aneuploidy)

Structural Chromosomal Abnormalities

• Deletions: cri-du-chat syndrome (5p– deletion → cat-like cry in infants, ID); DiGeorge syndrome (22q11.2 deletion → conotruncal heart defects, immunodeficiency, hypoparathyroidism)
• Inversions: Paracentric (doesn't include centromere; dicentric chromatids if crossing over within inversion); Pericentric (includes centromere)
• Translocations: Reciprocal (exchange between two non-homologous chromosomes); Robertsonian (fusion of two acrocentric chromosomes at centromere — 13, 14, 15, 21, 22; carrier has 45 chromosomes; risk of Down syndrome in offspring)
• Fragile sites: cytogenetically visible constrictions; Fragile X: FRAXA — CGG repeat (>200 → FMR1 methylation → silencing → loss of FMRP → X-linked intellectual disability)

Mutation and Mutagenesis

• Point mutations: Transition (purine→purine or pyrimidine→pyrimidine: A→G, C→T); Transversion (purine↔pyrimidine: A→C, G→T); Missense (codon → different AA — conservative if similar AA; non-conservative if different); Nonsense (codon → stop codon); Silent/synonymous (codon → same AA — wobble)
• Frameshift mutations: insertion or deletion of non-multiples of 3 → shift reading frame → garbled protein to next stop codon
• Trinucleotide repeat expansion: CGG in Fragile X (>200 repeats); CAG in Huntington disease (>36 repeats → polyQ → aggregates); CTG in myotonic dystrophy; anticipation — expansion increases in successive generations
• Physical mutagens: UV (pyrimidine dimers); ionising radiation (X-rays, γ-rays → DSBs, depurination); chemical mutagens: alkylating agents (EMS, MMS), base analogues (5-BU, 2-AP), intercalating agents (acridine orange, ethidium bromide → frameshift), oxidising agents (ROS)
• Ames test: tests chemical mutagenicity using His⁻ Salmonella typhimurium → if chemical mutates bacteria → growth on His⁻ medium → mutagenic (also often carcinogenic); includes rat liver extract (S9) to include metabolic activation

8.5 Recombinant DNA Technology

• Restriction enzymes (Type II): recognise specific palindromic sequences (4–8 bp); cut both strands → blunt or sticky ends; EcoRI (GAATTC → 5' AATT overhang); HindIII (AAGCTT → 5' AGCT); BamHI (GGATCC)
• Vectors: Plasmid (small circular DNA; ori + antibiotic resistance gene + MCS; up to ~10 kb insert); Bacteriophage (λ phage — 10–20 kb; M13 for ssDNA); Cosmid (plasmid + cos sites of λ; 35–45 kb); YAC (Yeast Artificial Chromosome; large inserts 200–1000 kb — used for Human Genome Project); BAC (Bacterial AC; 100–300 kb; more stable than YAC); Virus vectors (retrovirus, lentivirus, adenovirus, AAV for gene therapy)
• cDNA library: mRNA → reverse transcriptase → cDNA; represents expressed genes only (no introns); useful for eukaryotic gene expression in bacteria
• Genomic library: total genomic DNA cut with restriction enzyme → all sequences including introns and non-coding regions; complete but contains much non-coding DNA
• Polymerase Chain Reaction (PCR): in vitro DNA amplification; Denaturation (94°C) → Annealing (50–65°C, primers bind) → Extension (72°C, Taq polymerase extends 3'→5'); exponential amplification (2ⁿ copies after n cycles); applications: diagnostics (PCR of HIV, COVID-19), genotyping, forensics, quantitative PCR (qPCR/RT-PCR — measure gene expression)
• DNA sequencing: Sanger (dideoxy chain termination — ddNTPs stop extension; gel or capillary electrophoresis; gold standard but slow); Next-Generation Sequencing (NGS — Illumina: bridge amplification → sequencing by synthesis → massive parallel; Nanopore: real-time single molecule; long reads); WGS (whole genome), WES (whole exome), RNA-seq, ChIP-seq, ATAC-seq

CRISPR-Cas9 Genome Editing

• Cas9 (CRISPR-associated protein 9): endonuclease from Streptococcus pyogenes (SpCas9); guided by sgRNA (single guide RNA = crRNA + tracrRNA); recognises PAM (Protospacer Adjacent Motif — NGG for SpCas9); creates blunt DSB 3 bp upstream of PAM
• Repair outcomes: NHEJ (error-prone → indels → gene knockout); HDR (homology-directed repair with donor template → precise edits — knockin, correction)
• Variants: Base editors (CBE — C to T; ABE — A to G; no DSB); Prime editing (pegRNA — search and replace, no DSB); CRISPRi (dCas9 + KRAB → gene repression); CRISPRa (dCas9 + VPR → gene activation); Cas12a (TTTN PAM; staggered cuts); Cas13 (RNA-targeting)
• Applications: gene therapy (haemoglobinopathies — Casgevy for sickle cell/thalassaemia, FDA approved 2023); cancer immunotherapy (CAR-T editing); model organisms; agriculture (disease-resistant crops)
• Nobel Prize: Jennifer Doudna and Emmanuelle Charpentier (2020 Chemistry Nobel) for CRISPR-Cas9 development

Transgenics and Cloning

• Transgenic organisms: contain foreign gene (transgene) stably integrated in genome; Transgenic mice (pronuclear injection or ES cell → knockout; Cre-lox — conditional knockout); Transgenic plants (Agrobacterium tumefaciens Ti plasmid; biolistics/gene gun); Applications: Bt crops (Bacillus thuringiensis Cry toxin → insect resistance), Roundup-Ready (herbicide tolerance — EPSPS), Golden Rice (β-carotene biosynthesis genes — VAD intervention)
• Gene therapy: introducing functional gene to correct deficiency; Ex vivo (modify cells outside body, reinfuse — ADA-SCID, CAR-T); In vivo (vector directly into patient — Luxturna for RPE65 retinal dystrophy, Zolgensma for SMA); challenges: immune response, off-target integration, expression duration
• Genomics: Human Genome Project (completed 2003; 3.2 billion bp; ~20,000–25,000 protein-coding genes; only 1.5% codes for protein; 45% repetitive sequences); Personalised medicine; GWAS (genome-wide association studies)
• Proteomics: study of all proteins in cell/tissue; 2D gel electrophoresis + mass spectrometry; protein-protein interactions (Y2H, co-IP, BiFC); protein structure prediction (AlphaFold — AI-based, predicted structures of >200 million proteins)

9.1 Three Domain System and Five Kingdom Classification

• Three domain system (Woese, 1990 — based on 16S/18S rRNA sequences): Bacteria, Archaea, Eukarya; Archaea are more closely related to Eukarya than to Bacteria; supported by phylogenetic analysis, molecular data
• Five Kingdom classification (Whittaker, 1969): Monera (prokaryotes), Protista, Fungi, Plantae, Animalia; based on cell type, body organisation, nutrition, reproduction
• Two Kingdom (Linnaeus — Plantae and Animalia) → problematic for fungi, algae, bacteria

Viruses

• Non-living (no metabolism, no reproduction without host); contain either DNA or RNA (not both); obligate intracellular parasites; classified by nucleic acid type, strandedness, replication strategy (Baltimore classification — 7 groups)
• Structure: capsid (protein coat); nucleic acid; envelope (in some — from host membrane; phospholipid bilayer with viral glycoproteins); naked viruses (no envelope)
• Bacteriophage (phage): viruses infecting bacteria; lytic cycle (replication → cell lysis → burst); lysogenic cycle (phage genome integrates as prophage → replicates with host → can be induced → lytic cycle)
• Animal viruses: Adsorption → Penetration → Uncoating → Replication → Assembly → Release; RNA viruses: +ssRNA (picornavirus, coronavirus, flavivirus — directly translated); –ssRNA (rhabdovirus, orthomyxovirus — need RNA-dependent RNA polymerase first); dsRNA (rotavirus); retroviruses (HIV — reverse transcriptase → dsDNA integrates as provirus)
• HIV: envelope with gp120 (binds CD4 + CCR5 coreceptor) and gp41 (fusion); core: two ssRNA molecules + reverse transcriptase + integrase + protease; cycle: binding → fusion → RT → integration (provirus) → transcription → assembly → budding → maturation; HAART (highly active antiretroviral therapy) targets RT, protease, integrase, fusion
• Prions: infectious protein (PrPˢᶜ — misfolded form of normal PrPᶜ); cause spongiform encephalopathies: Scrapie (sheep), BSE (bovine), CJD (Creutzfeldt-Jakob — human), kuru (ritual cannibalism); no nucleic acid; templated misfolding; Stanley Prusiner (Nobel 1997)

Bacteria

• Structure: peptidoglycan cell wall; no membrane-bound nucleus; 70S ribosomes; circular chromosome; plasmids; pili (attachment, conjugation); flagella (motility — H antigen); capsule (virulence, anti-phagocytic)
• Gram staining: Gram-positive (thick peptidoglycan — retains crystal violet → purple; e.g., Staphylococcus, Streptococcus, Bacillus, Clostridium); Gram-negative (thin PG + outer membrane with LPS — decolorised by acetone → red with safranin; e.g., E. coli, Salmonella, Pseudomonas, Neisseria); outer membrane is LPS (lipopolysaccharide) — endotoxin
• Bacterial gene transfer: Transformation (uptake of naked DNA — basis of recombinant DNA technology; Griffith's experiment); Transduction (bacteriophage transfers DNA — generalised or specialised); Conjugation (cell-cell contact via pilus → plasmid/chromosome transfer — F plasmid; F⁺ → F⁻; Hfr strains — chromosomal gene transfer)
• Antibiotic resistance mechanisms: β-lactamase (enzyme — destroys penicillin/ampicillin/cephalosporin); Modified targets (PBP2a in MRSA — penicillin-binding protein); Efflux pumps (pump antibiotic out); Reduced permeability (modified porins); Enzymatic modification (aminoglycoside resistance)
• Biofilm: bacterial community encased in extracellular polysaccharide matrix; attached to surfaces; antibiotic-resistant (up to 1000× more resistant); chronic infections; dental plaque, cystic fibrosis lung, prosthetic device infections; quorum sensing regulates biofilm formation

Fungi

• Heterotrophic (absorptive nutrition); cell wall of chitin; ergosterol in membrane (target for antifungals — amphotericin B, azoles); dikaryotic phase in Basidiomycetes and Ascomycetes
• Classification: Zygomycetes (Rhizopus — bread mould; zygospore); Ascomycetes (largest group — yeast, Aspergillus, Penicillium, morels, truffles; sexual spores in asci); Basidiomycetes (mushrooms, rusts, smuts; sexual spores on basidia)
• Mycorrhizae: mutualistic association between fungi and plant roots; Ectomycorrhizae (sheath around roots — forest trees); Endomycorrhizae/Arbuscular mycorrhizae (hyphae inside root cells — 80% of plants); benefit: ↑ P and water uptake; pathogen resistance
• Medical importance: Candida albicans (oral/vaginal thrush — dimorphic); Aspergillus fumigatus (invasive aspergillosis in immunocompromised); Cryptococcus neoformans (meningitis in AIDS — India ink capsule); Dermatophytes (Tinea — ringworm, athlete's foot — Trichophyton, Microsporum, Epidermophyton)

Protists and Algae

• Diverse eukaryotic group — polyphyletic; includes photosynthetic (algae) and heterotrophic (protozoa, slime moulds) forms
• Algae: chlorophyll a in all; additional pigments vary; Chlorophyta (green algae — Chlamydomonas, Spirogyra, Volvox; closest relatives of land plants); Phaeophyta (brown algae — Fucus, Laminaria/kelp; algin); Rhodophyta (red algae — agar, carrageenan); Chrysophyta (diatoms — silica cell wall, diatomaceous earth; major primary producers); Dinoflagellates (red tides — paralytic shellfish poisoning; bioluminescence)
• Protozoa: Sarcomastigophora (amoeba, Giardia, Trypanosoma); Apicomplexa (Plasmodium, Toxoplasma — apical complex for invasion); Ciliophora (Paramecium — conjugation; micronucleus + macronucleus)
• Plasmodium lifecycle: sporozoites (from mosquito salivary gland) → hepatocytes (liver stage — hypnozoite in P. vivax, P. ovale → relapse) → merozoites → RBCs → ring trophozoite → schizont → merozoites (rupture — fever) → gametocytes → Anopheles mosquito → sexual stage (zygote → ookinete → oocyst → sporozoites)

10.1 Levels of Organisation and Energy Flow

• Ecological levels: individual → population → community → ecosystem → biome → biosphere
• Trophic levels: Producers (autotrophs — plants, algae, cyanobacteria); Primary consumers (herbivores); Secondary consumers; Tertiary consumers; Decomposers (detritivores — fungi, bacteria — break down dead organic matter)
• Food web: interconnected food chains; provides stability (if one species declines, energy flow can continue via other pathways); keystone species — disproportionate effect on ecosystem (sea otter — controls sea urchin → protects kelp forest)
• Energy flow: unidirectional; only 10% of energy passes between trophic levels (10% rule — Lindeman 1942); 90% lost as heat, respiration, uneaten biomass; Ecological Efficiency = (energy at trophic level n+1 / energy at trophic level n) × 100%
• Pyramids: Pyramid of numbers (can be inverted — trees); Pyramid of biomass (usually upright; inverted in open ocean — phytoplankton small but rapidly reproducing); Pyramid of energy (always upright — fundamental relationship)
• Primary productivity: GPP (Gross PP — total photosynthesis); NPP = GPP – Plant respiration; Net Ecosystem Production (NEP) = GPP – total ecosystem respiration; NEP >0 = carbon sink; tropical rainforests and estuaries have highest NPP

10.2 Biogeochemical Cycles

Carbon Cycle

• Photosynthesis (CO₂ fixation) ↔ Respiration (CO₂ release); decomposition; combustion of fossil fuels (adding ~10 PgC/year — exceeds natural sinks); oceanic uptake (~2.5 PgC/year); terrestrial uptake (~3.5 PgC/year); net increase ~3.5 ppm CO₂/year in atmosphere
• Greenhouse effect: CO₂, CH₄, N₂O, H₂O, ozone absorb outgoing infrared radiation → warm atmosphere; global warming; CO₂ GWP = 1 (reference); CH₄ GWP = 21 (over 100 years); N₂O GWP = 310; CFCs/HFCs — highest GWP

Nitrogen Cycle

• N₂ → NH₃ (Nitrogen fixation: Rhizobium, Azotobacter, Cyanobacteria, Anabaena; lightning; industrial Haber-Bosch); NH₃/NH₄⁺ → NO₂⁻ → NO₃⁻ (Nitrification: Nitrosomonas, Nitrobacter); NO₃⁻ → N₂ (Denitrification: Pseudomonas, Thiobacillus — anaerobic); organic N → NH₄⁺ (Ammonification by decomposers)
• Eutrophication: excessive nutrient input (N and P) → algal bloom → O₂ depletion (decomposers respire dead algae) → hypoxic dead zones → fish kills; example: Baltic Sea, Gulf of Mexico dead zone

Phosphorus Cycle

• No atmospheric reservoir (unlike C and N); weathering of phosphate rocks → dissolved phosphate → taken up by plants → food web → decomposition → soil; long-term storage in sedimentary rock; slow cycle; P often limiting nutrient in freshwater ecosystems

10.3 Population Ecology

• Population parameters: size (N), density, birth rate (b), death rate (d), immigration (i), emigration (e); dN/dt = (b + i) – (d + e)
• Exponential growth: dN/dt = rN; r = intrinsic rate of natural increase (b – d); occurs when resources unlimited; J-shaped curve; characteristic of invasive species, bacteria in favourable conditions
• Logistic growth: dN/dt = rN(K–N)/K; K = carrying capacity; S-shaped (sigmoidal) curve; population stabilises at K; density-dependent factors increase at high density (food competition, predation, disease, stress)
• Life history strategies: r-strategists (high r, many small offspring, little parental care, short lifespan — insects, weeds, mice; early colonisers); K-strategists (low r, few large offspring, extensive parental care, long lifespan — elephants, humans, trees; stable environments)
• Age structure: pre-reproductive, reproductive, post-reproductive; growing populations have large pre-reproductive cohort; declining populations have more post-reproductive; demographic transition
• Survivorship curves: Type I (high juvenile survival, most die of old age — humans, elephants); Type II (constant mortality across all ages — birds, many lizards); Type III (high early mortality, few survivors reproduce — oak trees, fish, most invertebrates)

10.4 Community Ecology and Succession

• Species interactions: Mutualism (+/+); Commensalism (+/0); Amensalism (–/0); Predation (+/–); Parasitism (+/–); Competition (–/–)
• Competitive Exclusion Principle (Gause's Law): two species competing for identical resources cannot coexist at stable equilibrium; one will be excluded; BUT niche differentiation allows coexistence
• Resource partitioning: sympatric species divide resources to avoid competition; Darwin's finches (beak size and food type); warblers (different foraging zones in spruce trees — MacArthur)
• Ecological niche: fundamental niche (physiological tolerance range — no competitors); realised niche (actual use in presence of competitors — smaller); hypervolume model (Hutchinson) — n-dimensional
• Predator-Prey dynamics: Lotka-Volterra equations; coupled oscillations (prey peaks → predator increases → prey declines → predator declines → prey recovers); Isle Royale (wolves and moose — classic example); top-down control (trophic cascade)
• Succession: Primary succession (bare substrate — no soil; e.g., bare rock, volcanic island — pioneer species: lichens, mosses → soil formation → grasses → shrubs → climax); Secondary succession (disturbed but soil intact — faster; abandoned field, fire, logging); Climax community (stable end state — debated; Clements vs Gleason view)

10.5 Biodiversity and Conservation

• Biodiversity levels: Genetic diversity (within species); Species diversity (alpha diversity — within habitat; beta diversity — between habitats; gamma diversity — overall landscape); Ecosystem diversity
• Biodiversity indices: Species richness (S); Shannon-Weiner diversity index H' = –Σ(pᵢ ln pᵢ); Simpson's index (D = Σnᵢ(nᵢ–1)/N(N–1)); Evenness (E = H'/H'max)
• Global biodiversity hotspots: 36 hotspots; must have ≥1500 endemic vascular plants AND ≥70% original habitat lost; India: Western Ghats + Sri Lanka; Eastern Himalayas; Indo-Burma
• IUCN Red List categories: Extinct (EX); Extinct in Wild (EW); Critically Endangered (CR — 80% decline in 10 years or 3 generations); Endangered (EN — 50% decline); Vulnerable (VU — 30% decline); Near Threatened (NT); Least Concern (LC)
• Major threats to biodiversity (HIPPO mnemonic): Habitat loss (most important — deforestation, agriculture, urban expansion); Invasive species; Pollution; Population (human); Overexploitation (hunting, poaching, overfishing)
• Conservation strategies: In situ (within natural habitat — national parks, sanctuaries, biosphere reserves, community reserves, Conservation Reserves); Ex situ (zoos, gene banks, seed banks — Svalbard Global Seed Vault; botanical gardens; cryopreservation)
• Protected areas in India: 106 National Parks; 567 Wildlife Sanctuaries; 18 Biosphere Reserves (12 in UNESCO MAB network); Project Tiger (1973 — now 54 tiger reserves; tiger population: ~3,167 in 2022); Project Elephant (1992); Operation Crocodile; Saving the Great Indian Bustard
• International agreements: CBD (Convention on Biological Diversity — 1992 Rio Earth Summit); CITES (Convention on International Trade in Endangered Species); RAMSAR (wetlands); IUCN; CMS (Migratory Species)

10.6 Environmental Issues

• Pollution types: Air (SO₂, NOₓ — acid rain; CO, PM₂.₅, PM₁₀, VOCs, O₃ — smog; AQI in India; NAAQS); Water (BOD — Biochemical Oxygen Demand; COD; heavy metals — Hg, Pb, Cd, As; pesticides; eutrophication); Soil (heavy metals, industrial effluents, plastic); Noise (decibels; WHO: <40 dB night outdoors, <70 dB workday); Radioactive (uranium mining, nuclear power, medical waste)
• Ozone depletion: Stratospheric ozone (O₃) absorbs UV-B and UV-C; CFCs (chlorofluorocarbons) → Cl• radicals → catalytically destroy O₃ (Cl• + O₃ → ClO + O₂; ClO + O → Cl• + O₂; one Cl• can destroy 100,000 O₃ molecules); Montreal Protocol (1987) — phase-out of CFCs; ozone hole recovery expected by 2065
• Climate change: ↑ CO₂ (from 280 ppm pre-industrial → 420 ppm 2023); global average temperature ↑1.1°C; consequences: sea level rise, glacier melt, extreme weather events, coral bleaching, range shifts of species, phenological changes; Paris Agreement (2015) — limit warming to 1.5°C above pre-industrial; India: NDC targets, renewable energy, Panchamrit commitments (COP26)
• Phytoremediation: use of plants to remove/degrade/detoxify pollutants from soil and water; Hyperaccumulators (Thlaspi for Zn/Cd; Brassica for Pb; Pteris fern for As); mechanisms: phytoextraction (plants take up metal in biomass), phytostabilisation, rhizofiltration, phytodegradation
• Bioremediation: use of microorganisms to degrade pollutants; Pseudomonas putida (degrades petroleum hydrocarbons — first patented organism: Ananda Chakraborty, 1980); mycoremediation (fungi — Phanerochaete chrysosporium degrades lignin and toxic organics); biostimulation (add nutrients to stimulate native microbes); bioaugmentation (add specific microbes)

11.1 Origin of Life

• Chemical evolution: Oparin-Haldane hypothesis (1920s) — primitive Earth had reducing atmosphere (H₂, NH₃, CH₄, H₂O); UV light and lightning provided energy → synthesis of organic molecules → 'primordial soup'
• Miller-Urey experiment (1953): simulated primitive Earth conditions → produced amino acids, sugars, organic acids; demonstrated abiotic synthesis of organic molecules
• RNA World hypothesis: RNA both carries information (like DNA) and has catalytic activity (ribozymes); RNA preceded DNA and proteins; ribosomes still use RNA (23S rRNA) for peptide bond formation — fossil of RNA world
• LUCA (Last Universal Common Ancestor): ~3.5–4 billion years ago; had ribosomes, genetic code, cell membrane; ancestors of Bacteria, Archaea, Eukarya
• Earliest life evidence: stromatolites (3.5 Bya — microbial mats); microfossils (Apex Chert, Australia); molecular fossils; Cyanobacteria responsible for Great Oxidation Event (~2.4 Bya) — O₂ accumulation in atmosphere

11.2 Theories of Evolution

• Lamarckism (Jean-Baptiste Lamarck, 1809): inheritance of acquired characteristics; use and disuse; directed towards greater complexity; disproved (Weismann's experiment — cutting tails of mice for many generations; molecular genetics)
• Darwinism (Charles Darwin, 1859 — On the Origin of Species): Natural Selection; variation exists in populations; individuals with advantageous traits survive and reproduce more (differential fitness); traits heritable → gradual change over generations
• Neo-Darwinism/Modern Synthesis (1930s–40s): integration of Darwinian natural selection with Mendelian genetics; Dobzhansky, Mayr, Huxley, Simpson; population genetics framework (Hardy-Weinberg)

Mechanisms of Evolution

Hardy-Weinberg Equilibrium

• In an ideal large random-mating population with no mutation, no migration, no selection → allele and genotype frequencies remain constant
• HWE equations: p + q = 1 (allele frequencies); p² + 2pq + q² = 1 (genotype frequencies — AA, Aa, aa)
• Five assumptions of HWE (if ANY violated → evolution occurs): Random mating; No selection; No mutation; No gene flow; Very large population size
• Application: detect non-random mating (inbreeding — excess homozygotes); estimate carrier frequency (cystic fibrosis — q² = 1/2500, q = 1/50, 2pq ≈ 1/25 carriers)

11.3 Evolutionary Patterns and Speciation

• Species concepts: Biological Species Concept (Mayr — interbreeding natural populations reproductively isolated from others; problematic for fossils, asexual, ring species); Phylogenetic/Cladistic; Ecological; Morphological/Typological
• Speciation types: Allopatric (geographic isolation → independent evolution → reproductive isolation; most common; mountain ranges, rivers, ocean → vicariance; Galapagos finches — island colonisation); Parapatric (adjacent populations; partial barrier; along environmental gradient); Sympatric (in same geographic area — polyploidy in plants, host switching in insects, assortative mating)
• Reproductive isolating mechanisms: Pre-zygotic (habitat, temporal/seasonal, behavioural, mechanical, gametic isolation — prevent mating or fertilisation); Post-zygotic (hybrid inviability, hybrid sterility — mule from horse × donkey, hybrid breakdown)
• Evolutionary patterns: Divergent evolution (common ancestor → different forms; homologous structures — forelimbs of mammals); Convergent evolution (unrelated species → similar form due to similar selection pressure; analogous structures — wings of birds, bats, insects, pterosaurs; eye in vertebrates and octopus); Parallel evolution (related species independently evolve similar traits)
• Mimicry: Batesian (harmless species mimics harmful/toxic species — hoverfly mimics wasp); Müllerian (two or more harmful species share warning coloration — monarch and viceroy butterflies)
• Co-evolution: reciprocal evolutionary change; parasite-host; predator-prey; flowering plants and pollinators (orchid-moth; fig-fig wasp); Red Queen hypothesis — constant evolutionary arms race between species

11.4 Human Evolution

• Primate phylogeny: Order Primates → Prosimii (lemurs, lorises, tarsiers) + Anthropoidea (New World monkeys, Old World monkeys, Apes + Humans)
• Great apes: Pongidae — Orangutan (Pongo), Gorilla, Chimpanzee (Pan — Pan troglodytes and Pan paniscus/bonobos); Humans most closely related to chimpanzees (~98% DNA identity)
• Human fossil record:

• Mitochondrial Eve: most recent common matrilineal ancestor of all living humans; ~150,000–200,000 years ago in Africa; traced via mtDNA (maternal inheritance only); Y-chromosome Adam (~200,000–300,000 ya — most recent common patrilineal ancestor)
• Out of Africa theory: H. sapiens evolved in Africa → single migration out ~60,000–70,000 years ago → spread globally; supported by greatest genetic diversity in Africa (more time for divergence)

11.5 Animal Behaviour (Ethology)

• Innate (fixed) behaviour: genetically programmed; not learned; same in all individuals of species regardless of experience
• Fixed Action Patterns (FAPs): stereotyped, species-specific motor sequences triggered by sign stimulus (releaser); Lorenz and Tinbergen; egg-rolling by Greylag goose; gaping of nestlings; stickleback aggression to red belly
• Sign stimulus and supernormal stimuli: sign stimulus is minimum stimulus needed to trigger FAP; supernormal stimulus is exaggerated version → stronger response (giant egg preferred over own egg by oystercatcher; human fondness for baby faces)
• Tinbergen's four questions (levels of analysis): Causation (mechanism/how?), Ontogeny (development — how does it develop?), Function/Adaptation (why — survival value?), Phylogeny (evolution — how did it evolve?)

Learning Types

Social Behaviour and Communication

• Waggle dance of honeybees (Karl von Frisch — Nobel 1973): worker bee communicates distance and direction of food source; round dance (<100m); waggle dance (>100m); angle of dance from vertical = angle of food from sun; duration of waggle run = distance; figure-eight pattern
• Altruism and kin selection: altruistic behaviour reduces actor's fitness but benefits recipient; Hamilton's Rule: rB > C (r = relatedness coefficient, B = benefit to recipient, C = cost to actor); kin selection — helping relatives passes shared genes; Inclusive fitness = direct fitness + indirect fitness (via helping relatives)
• Eusociality: highest level of social organisation; reproductive division of labour; overlapping generations; cooperative brood care; Hymenoptera (bees, wasps, ants — haplo-diploid: workers r=0.75 to sisters); naked mole rat (only eusocial mammal)
• Reciprocal altruism (Robert Trivers): altruism extended to non-relatives when favour likely to be returned; 'you scratch my back, I'll scratch yours'; requires recognition, memory, repeated interactions; vampire bats blood sharing
• Mating systems: Monogamy (one mate; high parental certainty; many birds); Polygyny (one male, multiple females; sexual dimorphism in males; lions, red deer); Polyandry (one female, multiple males; reversed sexual dimorphism; jacanas, seahorses); Promiscuity (no pair bond)
• Parental investment theory (Robert Trivers): the sex that invests more in offspring (usually female) will be choosier in mate selection; the less investing sex will compete for access
• Pheromones: chemical signals between conspecifics; primer pheromones (alter physiology — queen bee pheromone prevents worker ovary development); releaser pheromones (immediate behaviour — sex attractants in moths, alarm in ants); Bombykol (silk moth sex pheromone — first pheromone identified)

12.1 Microbial Fermentation and Biotechnology

• Fermentation (industrial): microbial production of useful products; batch, fed-batch, or continuous culture; industrial fermentors (bioreactors) — controlled temperature, pH, aeration, agitation
• Products of microbial fermentation: Ethanol (Saccharomyces cerevisiae — glucose → pyruvate → ethanol + CO₂); Lactic acid (Lactobacillus — yogurt, cheese, sauerkraut, silage); Citric acid (Aspergillus niger); Glutamate (Corynebacterium glutamicum — monosodium glutamate, MSG); Lysine (C. glutamicum — animal feed supplement); Penicillin (Penicillium chrysogenum); Streptomycin (Streptomyces griseus); Insulin (E. coli recombinant); Erythropoietin (CHO cells); tPA
• Antibiotic production: Secondary metabolites from fungi (Penicillium, Cephalosporium) and actinomycetes (Streptomyces — streptomycin, erythromycin, tetracycline, vancomycin, amphotericin B); produced in stationary phase
• SCP (Single Cell Protein): microbial biomass as protein source; Spirulina (cyanobacterium — 60–70% protein); Quorn (Fusarium venenatum — mycoprotein); advantages: rapid growth, use of waste substrates, high protein content

12.2 Immunology Applications

• Vaccines: Active immunisation — stimulate immune memory; Live attenuated (BCG, MMR, oral polio, varicella — more effective, may revert); Killed/inactivated (IPV, hepatitis A, flu injectable — safer); Subunit/recombinant (hepatitis B surface antigen from yeast, acellular pertussis, HPV — VLP); Toxoid (tetanus, diphtheria — inactivated toxin); DNA vaccines; mRNA vaccines (COVID-19 — Pfizer/Moderna — BNT162b2, mRNA-1273)
• Passive immunisation: injection of antibodies; immediate protection, no memory; antivenom (anti-snake venom antibodies); rabies post-exposure; maternal IgG across placenta (first 6 months of life protection)
• ELISA (Enzyme-Linked ImmunoSorbent Assay): Direct, Indirect, Sandwich, Competitive formats; diagnoses HIV (anti-HIV Ab), hepatitis B/C, pregnancy test (hCG), allergy testing, food contaminants; high-throughput; uses enzyme-linked secondary Ab → colorimetric reaction → measured by spectrophotometry
• Western Blot (Immunoblotting): SDS-PAGE → transfer to nitrocellulose membrane → block with milk/BSA → primary Ab → secondary Ab (enzyme/fluorescent) → detection; confirms HIV diagnosis; detects specific proteins; Prion disease diagnosis (PrPˢᶜ); protein size determination
• Flow Cytometry and Immunofluorescence: fluorescence-labelled Abs + cells in flow → laser excites → detectors measure scatter and fluorescence; counts and characterises cells; CD4 count in HIV; cancer immunophenotyping; FACS (fluorescence-activated cell sorting)
• Monoclonal antibodies (mAbs): Köhler and Milstein (Nobel 1984); B cell (immunised mouse) + myeloma cell → hybridoma → clones produce single specificity Ab; Therapeutic mAbs: Trastuzumab/Herceptin (anti-HER2 — breast cancer); Rituximab (anti-CD20 — lymphoma); Adalimumab/Humira (anti-TNFα — rheumatoid arthritis — best-selling drug); Checkpoint inhibitors (anti-PD1 — pembrolizumab/nivolumab; anti-CTLA4 — ipilimumab) — cancer immunotherapy

12.3 Agricultural and Environmental Biotechnology

• Plant tissue culture (PTC): totipotency (Haberlandt's concept — 1902; first demonstrated by Steward with carrot cells, 1958); MS medium (Murashige and Skoog — inorganic salts + vitamins + sucrose + agar); phytohormone balance determines differentiation (high auxin:cytokinin → root; low A:C → shoot; equal → callus)
• Applications of PTC: Micropropagation (clonal propagation of elite plants — orchids, banana, potato); Somatic hybridisation (protoplast fusion — pomato = potato + tomato; not commercially viable but demonstrated); Haploid production (anther culture → doubled haploids → homozygous lines in one generation); Somaclonal variation; Production of secondary metabolites (taxol from Taxus bark vs cell culture)
• Biofertilisers: living organisms that supply nutrients; Rhizobium (N fixation in legumes); Azotobacter, Azospirillum (free-living N fixers); Cyanobacteria (Anabaena — rice fields); Mycorrhizae (P solubilisation, mineral uptake); Phosphate-solubilising bacteria (Bacillus, Pseudomonas); advantages: cheaper, eco-friendly, improve soil health
• Biopesticides: biological pest control; Bt (Bacillus thuringiensis) — Cry proteins (δ-endotoxins) → bind gut epithelium of insects → pore formation → ion imbalance → death; specific to lepidoptera, diptera, coleoptera; used as spray or transgenic crops (Bt cotton, Bt brinjal); NPV (Nuclear Polyhedrosis Virus) for caterpillars; Trichoderma (biocontrol against soil fungi)
• Genetically Modified (GM) crops: herbicide tolerance (Roundup Ready — EPSPS*); insect resistance (Bt crops); disease resistance; enhanced nutrition (Golden Rice — psy + crtI genes from daffodil/bacteria → β-carotene); drought tolerance; extended shelf life (Flavr Savr tomato — anti-sense ACC synthase); India approved Bt cotton (2002) and is considering Bt brinjal, GM mustard
• Metagenomics: study of genetic material from environmental samples (soil, gut, ocean) without culturing; reveals unculturable organisms (99% of environmental microbes cannot be cultured in lab); Human Microbiome Project; Earth Microbiome Project; functional metagenomics for novel enzymes

12.4 Bioresource and Biodiversity Utilisation

• Bioprospecting: systematic search for organisms and molecules that may be useful commercially; often in biodiversity hotspots; concerns about biopiracy (unauthorised use of traditional knowledge — neem, turmeric patent controversies)
• Access and Benefit Sharing (ABS): Nagoya Protocol (2010 — supplementary to CBD); ensures fair sharing of benefits from use of genetic resources with country of origin; India: Biological Diversity Act 2002; National Biodiversity Authority (NBA)
• Traditional knowledge: indigenous communities' knowledge of medicinal plants, agricultural practices; documentation in TKDL (Traditional Knowledge Digital Library — India, to defend against biopiracy); Neem (biopesticide, antifungal — patent challenged); Turmeric (wound healing — patent revoked); Basmati rice (GI — Geographical Indication)
• Blue biotechnology: marine organisms for biotechnology; ω-conotoxins (cone snail — chronic pain treatment); Taq polymerase (Thermus aquaticus — hot spring bacterium); deep-sea microbes (extremophiles — extremozymes for industrial use)

13.1 Microscopy

13.2 Separation Techniques

Centrifugation

• Separation based on sedimentation rate (size and density); rate = sedimentation coefficient (Svedberg, S units)
• Differential centrifugation: sequential centrifugation at increasing speeds; 600g (nuclei, unbroken cells) → 10,000g (mitochondria, chloroplasts, lysosomes) → 100,000g (microsomes — ER, Golgi) → supernatant (cytosol)
• Density gradient centrifugation: Sucrose gradient (rate-zonal — separate by size, shape); CsCl gradient (equilibrium/isopycnic — separate by density; used in Meselson-Stahl experiment to separate ¹⁵N/¹⁴N DNA; separate DNA from RNA and protein)
• Ultracentrifugation: up to 500,000g; Sedimentation of ribosomes (70S prokaryotic: 50S + 30S; 80S eukaryotic: 60S + 40S); macromolecule characterisation

Chromatography

Electrophoresis

• Agarose gel electrophoresis: DNA/RNA separation by size; negatively charged nucleic acids migrate toward positive pole; smaller molecules move faster; visualised with ethidium bromide (EtBr) or SYBR Green + UV light; DNA ladder as size marker; applications: PCR product analysis, RFLP, Southern blot preparation
• SDS-PAGE (SDS-Polyacrylamide Gel Electrophoresis): SDS denatures proteins + imparts uniform negative charge per unit mass → separation by size only; reducing conditions break disulfide bonds; Staining: Coomassie blue (general), silver stain (high sensitivity), SYPRO Ruby (fluorescent)
• 2D gel electrophoresis: IEF (Isoelectric Focusing — first dimension, separates by pI) + SDS-PAGE (second dimension, separates by size) → each spot = one protein; map of entire proteome; limitations: hydrophobic proteins, dynamic range
• Capillary electrophoresis: high voltage in thin capillary; high resolution; automated; Sanger sequencing readout
• Southern blot: DNA → gel → transfer to membrane → hybridise with probe → detect; detects specific DNA sequences/mutations; RFLP analysis
• Northern blot: RNA → gel → membrane → probe → detects mRNA; size and abundance of specific mRNA
• Western blot: protein → SDS-PAGE → membrane → primary Ab → secondary Ab-enzyme → ECL detection; detects specific proteins; see immunology applications above

13.3 Molecular Biology Techniques

• RT-PCR (Reverse Transcription PCR): RNA → cDNA (reverse transcriptase) → PCR amplification; detects RNA viruses (HIV, COVID-19); measures gene expression (semi-quantitative)
• qPCR (Quantitative/Real-Time PCR): fluorescent signal (SYBR Green or TaqMan probe) measured each cycle; quantifies DNA/RNA in real time; Cᵧ value (cycle at which fluorescence crosses threshold) → lower Cᵧ = more template; gene expression quantification (ΔΔCᵧ method)
• Gene expression analysis: Microarray (DNA chips — thousands of probes on chip; hybridise labelled cDNA from two conditions; red/green ratio = differential expression; older technology); RNA-seq (NGS-based; digital counting; wider dynamic range; detects novel transcripts, splice variants; replaced microarrays)
• ChIP-seq (Chromatin Immunoprecipitation + Sequencing): crosslink protein-DNA → shear chromatin → immunoprecipitate with Ab against specific protein (TF, histone modification) → reverse crosslinks → sequence → maps genomic binding sites; identifies enhancers, promoters, histone modification patterns
• ATAC-seq: Assay for Transposase-Accessible Chromatin — maps open chromatin regions; transposase cuts and inserts sequencing adaptors preferentially in accessible chromatin → identifies active regulatory elements
• Protein sequencing: Edman degradation (sequential removal of N-terminal amino acids — phenylisothiocyanate; determines sequence; automated; up to ~60 residues); Mass spectrometry (trypsin digestion → peptides → MALDI-TOF or ESI-MS/MS → de novo sequencing or database matching)
• X-ray crystallography: protein/nucleic acid crystals → X-ray diffraction → electron density map → atomic model; determined DNA structure (Franklin's photo 51); many enzyme structures; PDB (Protein Data Bank — 200,000+ structures)
• NMR spectroscopy: solution structure; dynamics; protein-ligand interactions; limited to smaller proteins (<50 kDa); useful for intrinsically disordered proteins
• Two-Hybrid System (Yeast Two-Hybrid, Y2H): detecting protein-protein interactions in vivo; bait protein fused to DNA-BD; prey protein fused to AD; interaction → reporter gene expression; limitations: false positives, requires nuclear localisation, can't study toxic interactions
• FRET (Förster Resonance Energy Transfer): distance between fluorophores <10 nm → energy transfer from donor to acceptor; detects molecular interactions, conformational changes in live cells; single-molecule FRET (sm-FRET)

13.4 Antibody-Based Detection Methods

• ELISA (Enzyme-Linked ImmunoSorbent Assay): see above; highly sensitive and specific; OD (optical density) measured at specific wavelength (450 nm for TMB substrate)
• Radioimmunoassay (RIA): uses radioactive isotope-labelled antigen (¹²⁵I); competitive binding; highly sensitive; now replaced by ELISA in most applications due to radiation concerns
• Immunofluorescence: Direct (fluorophore-labelled primary Ab) or Indirect (unlabelled primary + fluorophore-secondary — amplification); co-localisation studies; fixed (formaldehyde) or live cell imaging; confocal for 3D
• Immunoprecipitation (Co-IP): antibody + cell lysate → bead pull-down → wash → elute → western blot; detects protein-protein interactions in native conditions; ChIP uses this principle with DNA

13.5 Animal and Cell Culture Models

• Cell culture: primary cells (directly from tissue — limited divisions); cell lines (transformed — immortal; HeLa — first human cell line, Henrietta Lacks 1951, HPV-18 oncogene; CHO cells — biopharmaceutical production; 3T3 fibroblasts; Jurkat — T cell line)
• Stem cell culture: ESC (embryonic stem cells — require feeder cells, LIF in mouse; 3D organoids); iPSC (reprogrammed; patient-specific; disease modelling); maintenance of pluripotency vs directed differentiation
• Model organisms: E. coli (gene regulation, molecular biology); Saccharomyces cerevisiae (cell cycle — Hartwell Nobel 2001; protein secretion); Caenorhabditis elegans (fixed cell lineage; apoptosis; RNAi; Brenner, Sulston, Horvitz Nobel 2002); Drosophila melanogaster (genetics — Morgan Nobel 1933; development — Nüsslein-Volhard/Wieschaus Nobel 1995; behaviour); Danio rerio (zebrafish — transparent embryo; vertebrate development; ENU mutagenesis screens); Mus musculus (mouse — gene knockout, transgenic; disease models); Arabidopsis thaliana (plant genomics; small genome; rapid life cycle; T-DNA insertion mutants)
• 3R Principle (animal experimentation): Replacement (use non-animal alternatives where possible); Reduction (use minimum number of animals); Refinement (minimise suffering, improve procedures); IAEC (Institutional Animal Ethics Committee) oversight required in India (CPCSEA regulation)

PYQ.1 Most Frequently Asked Topics

PYQ.2 Year-wise High-Value Questions (Selected)

QR.1 Nobel Prizes — Life Sciences (Must Know)

QR.2 Mnemonics

QR.3 Key Numbers and Values

QR.4 Molecular Biology Quick Reference

QR.5 Cell Signaling Receptors — Quick Reference

QR.6 Hormones and Control — Quick Reference

AW.1 Directive Words and Answer Structure

AW.2 High-Scoring Answer Templates

AW.3 Must-Draw Diagrams

AW.4 Exam-Day Strategy

• First 15 minutes: Read all questions carefully; mark your sure-shot 4 questions; plan which optional to attempt for 5th question; allocate time (20-mark questions: 25–30 min each; 10-mark: 12–15 min each)
• Diagram first strategy: For mechanism questions, draw the diagram FIRST before writing; it organises your thoughts and ensures you don't miss the diagram at the end
• Start with strongest question: Attempt the question you know best first; momentum and confidence; examiner sees your best work early
• Quantitative questions: Show ALL working; even if final answer is wrong, partial credit for correct steps; check dimensional analysis; state formula before applying values
• Keywords: Use technical terms correctly and precisely — examiners look for domain-specific vocabulary (e.g., 'Ku70/Ku80 heterodimer' not just 'proteins'; 'PCNA sliding clamp' not just 'processivity factor')
• Indian context bonus: Always add India-specific examples where possible (India's first test tube baby, Bt cotton controversies, Western Ghats biodiversity, NIN/NCBS research) — differentiates your answer
• End each answer clearly: A brief conclusion restating the main point; examiners often skim to the end

30-Day Revision Plan

AW.5 Interdisciplinary Connections for Bonus Marks

• Molecular biology + Medicine: Link BRCA1/2 mutations to cancer and CRISPR therapy; p53 dysfunction to tumour biology; telomerase to ageing and cancer
• Cell signaling + Disease: Oncogenic Ras → colorectal, lung, pancreatic cancer; EGFR mutations → lung cancer (gefitinib target); BCR-Abl → CML (imatinib/Gleevec first targeted therapy)
• Genetics + Ethical dimensions: Human genome editing (germline vs somatic); He Jiankui scandal (CCR5 editing in embryos); designer babies; genetic discrimination; Genetic Data Protection
• Ecology + Climate change: Range shifts of species northward; phenological mismatch (flowers early → pollinators not synchronised); coral bleaching (thermal stress → symbiotic algae expulsion); India: glacial retreat in Himalayas affecting agriculture
• Evolution + Antibiotic resistance: Classic example of natural selection in real time; MRSA in hospital settings; resistance to antimalarials (Plasmodium falciparum resistance to chloroquine, artemisinin); WHO priority pathogens
• Plant physiology + Agriculture: Dwarf wheat (Green Revolution — semi-dwarfing Rht genes — GA-insensitive DELLA proteins); photoperiodism in rice — IR8 (short-day) vs Basmati (photoperiod-sensitive); nitrogen fixation and biofertilisers
• Methods + Real-world application: PCR in COVID diagnosis (RT-PCR); CRISPR in sickle cell treatment (Casgevy — FDA 2023); monoclonal antibodies in cancer immunotherapy; flow cytometry in HIV monitoring (CD4 count)