Molecular Basis of Inheritance — Long Notes
Mendel's principles gave us the "what" of inheritance. The molecular basis gives us the "how": the chemical identity of the hereditary material, its structure, and how it is copied, expressed, and regulated.
1. The Genetic Material: Journey to DNA
1.1 Griffith (1928) — Transformation
Working with Streptococcus pneumoniae:
- Live S (smooth, virulent) → mice died.
- Live R (rough, non-virulent) → mice lived.
- Heat-killed S → mice lived.
- Heat-killed S + live R → mice died — some substance in dead S transformed R into virulent form. He called it the "transforming principle".
1.2 Avery, MacLeod & McCarty (1944) — Transforming Principle = DNA
They repeated Griffith's experiment, treating the heat-killed S extract with different enzymes:
- Digested proteins (protease) → still virulent.
- Digested RNA (RNase) → still virulent.
- Digested DNA (DNase) → no transformation.
Conclusion: DNA is the transforming principle.
1.3 Hershey & Chase (1952) — Confirmed with Phage
- Bacteriophage T2 infects E. coli.
- Two batches of phages: one grown with ³²P (labels DNA), other with ³⁵S (labels protein).
- After infection and blender-shearing, they measured which label entered bacteria.
- Only ³²P went in — DNA (not protein) is injected.
- Definitive proof: DNA is the genetic material.
2. DNA Structure — Watson & Crick (1953)
Built on:
- Chargaff's rules (A = T, G = C, A + G = T + C).
- X-ray diffraction photos by Rosalind Franklin & Wilkins.
2.1 The Double Helix
- Two polynucleotide strands wound as a right-handed double helix.
- Strands are antiparallel — one runs 5'→3', the other 3'→5'.
- Sugar-phosphate backbone on the outside; bases on the inside, paired by H-bonds:
- Adenine (A) – Thymine (T) with 2 H-bonds.
- Guanine (G) – Cytosine (C) with 3 H-bonds.
- Dimensions: diameter 2 nm; each base-pair rise 0.34 nm; 10 bp per turn; pitch 3.4 nm.
- Grooves: major and minor — where regulatory proteins bind.
2.2 Chemistry
- Nucleotide = phosphate + pentose + nitrogen base.
- In DNA sugar = deoxyribose; bases = A, G (purines), C, T (pyrimidines).
- In RNA sugar = ribose; T → U (uracil).
3. DNA Packaging
- Human diploid genome ≈ 3.1 × 10⁹ bp; total length ~2.2 m stretched. Must be packed into a nucleus < 10 μm.
- Prokaryotes — no true nucleus; DNA (as nucleoid) is negatively charged and organised with positively charged proteins into supercoiled loops.
- Eukaryotes — DNA wraps around histones (H2A, H2B, H3, H4) as an octamer (2 of each). Each nucleosome has ~200 bp of DNA. H1 histone binds between nucleosomes.
- Nucleosome + linker DNA = a "beads on a string" appearance.
- Further coiling → 30-nm fibre → looped domains → chromatin → chromosomes.
- Euchromatin — loosely packed, lightly stained, transcriptionally active.
- Heterochromatin — densely packed, darkly stained, transcriptionally inactive.
4. DNA Replication
DNA must copy itself faithfully before every cell division. Watson-Crick model implied "semi-conservative" replication — each daughter gets one old + one new strand.
4.1 Meselson & Stahl (1958)
- Grew E. coli for many generations in ¹⁵N medium — all DNA became heavy.
- Shifted to ¹⁴N medium.
- After one generation: DNA was intermediate density (hybrid).
- After two generations: 50% intermediate + 50% light.
- Only semi-conservative replication fits — confirmed unambiguously.
4.2 Mechanism (in E. coli)
- Initiation at the origin of replication (ori).
- Helicase unwinds the double helix → replication fork.
- Topoisomerase (gyrase) relieves supercoiling ahead.
- Primase lays down short RNA primers.
- DNA polymerase III adds nucleotides only in the 5'→3' direction.
- Leading strand: continuous synthesis toward the fork.
- Lagging strand: synthesised discontinuously, in short Okazaki fragments away from the fork.
- DNA polymerase I replaces RNA primers with DNA.
- DNA ligase seals the nicks between Okazaki fragments.
Replication is amazingly accurate (~1 error in 10⁹ bases) thanks to polymerase proof-reading + mismatch repair.
5. Transcription (DNA → RNA)
5.1 Basics
- Only one strand of the DNA (the template strand, 3'→5') is transcribed.
- The other strand — the coding (sense) strand — has the same sequence as the RNA (except T → U).
- Catalysed by RNA polymerase, which synthesises RNA 5'→3'.
5.2 Prokaryotic vs Eukaryotic Transcription
Prokaryotes:
- Single RNA polymerase makes mRNA, rRNA, tRNA.
- No nucleus → transcription and translation are coupled (happen simultaneously).
- Sigma factor helps polymerase find the promoter.
- Rho protein helps terminate.
Eukaryotes:
- Three RNA polymerases:
- RNA pol I — rRNA (except 5S).
- RNA pol II — mRNA precursor (hnRNA).
- RNA pol III — tRNA, 5S rRNA, snRNAs.
- Primary transcript (hnRNA) has introns (non-coding) and exons (coding).
- Processing before export:
- 5' Capping — 7-methyl guanosine added.
- 3' Tailing (polyadenylation) — ~200 A residues.
- Splicing — spliceosome removes introns and joins exons.
- Mature mRNA is exported to the cytoplasm for translation.
6. The Genetic Code
Deciphered by Nirenberg, Khorana, Holley, and others in the 1960s.
Properties
- Triplet — 3 bases (codon) = 1 amino acid.
- Degenerate — most amino acids have >1 codon.
- Unambiguous — each codon codes for one specific amino acid.
- Universal — same code across species (rare exceptions in mitochondria/some protists).
- Non-overlapping and comma-less — read in fixed frame, no gaps.
- Initiation codon — AUG (also codes for methionine).
- Termination codons — UAA, UAG, UGA (do not code for any amino acid).
7. tRNA — the Adapter
Predicted by Crick, discovered later.
- 2D structure: cloverleaf — with an anticodon loop and an amino-acid attachment site (3'-CCA).
- 3D structure: L-shape.
- Each tRNA is charged by a specific aminoacyl-tRNA synthetase — pairing the correct amino acid to the correct anticodon.
8. Translation
Occurs on ribosomes. Ribosome = 2 subunits (60S + 40S in eukaryotes; 50S + 30S in prokaryotes).
8.1 Stages
- Initiation — small subunit binds mRNA at AUG; initiator tRNA (with Met/fMet) enters the P-site; large subunit joins.
- Elongation — next aminoacyl-tRNA enters A-site → peptide bond forms (catalysed by 23S rRNA — a ribozyme) → tRNA in P-site moves to E-site and leaves → ribosome shifts one codon (translocation).
- Termination — release factor recognises stop codon → polypeptide released.
The polypeptide then folds into its 3D shape (sometimes with help of chaperones) and may undergo post-translational modifications.
9. Regulation of Gene Expression
Cells don't need all proteins all the time. Regulation occurs at multiple levels — for our syllabus, focus on prokaryotic transcriptional control via the lac operon.
The Lac Operon (Jacob & Monod, 1961)
In E. coli, three structural genes are transcribed together as a single mRNA:
- z — β-galactosidase (hydrolyses lactose → glucose + galactose).
- y — permease (imports lactose).
- a — transacetylase.
Upstream:
- p — promoter (RNA pol binds here).
- o — operator (repressor binding site).
- i — regulator gene, transcribed independently, produces the repressor protein.
Absent lactose: repressor binds the operator → blocks polymerase → no transcription. Present lactose: some lactose enters the cell → converted to allolactose → binds the repressor → repressor releases the operator → transcription ON.
This is negative regulation with an inducer — a classic on/off switch.
10. Human Genome Project (1990–2003)
- Massive international effort to sequence the entire human genome.
- Sequenced by two consortia (public + Celera).
Key findings
- ~3.1 × 10⁹ base pairs in the haploid genome.
- ~30,000 genes — far fewer than the original estimate of 1 lakh.
- ~2% of DNA codes for proteins — most is regulatory, structural, or non-coding.
- Chromosome 1 has the most genes (~2,968); Y has the fewest (~231).
- SNPs (single nucleotide polymorphisms) — millions, useful for tracing ancestry and disease predisposition.
- Two randomly selected humans are 99.9% identical at DNA level.
11. DNA Fingerprinting
Alec Jeffreys (1985) developed the technique.
- Based on VNTRs (Variable Number Tandem Repeats) — short DNA sequences repeated a variable number of times among individuals.
- Steps:
- Isolate DNA from sample (blood, hair, semen, saliva).
- Digest with restriction enzymes.
- Separate fragments by gel electrophoresis.
- Transfer to nylon membrane (Southern blot).
- Hybridise with a labelled VNTR probe.
- Detect by autoradiography — the resulting band pattern is the "fingerprint".
Applications: forensic identification, paternity testing, tracing pedigrees, wildlife conservation, and evolution studies.
Key take-aways
- DNA is the genetic material — proved by Avery et al. and Hershey–Chase.
- Structure = function: the antiparallel double helix with A-T, G-C pairing enables fidelity, replication, and coding.
- Central dogma: DNA → RNA → Protein, with reverse transcription in retroviruses.
- Prokaryotes use one RNA polymerase; eukaryotes use three plus extensive mRNA processing.
- The genetic code is triplet, degenerate, and (nearly) universal.
- Regulation examples: the lac operon is a classic negative-inducible system.
- The Human Genome Project and DNA fingerprinting brought molecular biology into medicine, forensics, and everyday life.
Molecular Basis of Inheritance — Long Notes
Mendel's principles gave us the "what" of inheritance. The molecular basis gives us the "how": the chemical identity of the hereditary material, its structure, and how it is copied, expressed, and regulated.
1. The Genetic Material: Journey to DNA
1.1 Griffith (1928) — Transformation
Working with Streptococcus pneumoniae:
- Live S (smooth, virulent) → mice died.
- Live R (rough, non-virulent) → mice lived.
- Heat-killed S → mice lived.
- Heat-killed S + live R → mice died — some substance in dead S transformed R into virulent form. He called it the "transforming principle".
1.2 Avery, MacLeod & McCarty (1944) — Transforming Principle = DNA
They repeated Griffith's experiment, treating the heat-killed S extract with different enzymes:
- Digested proteins (protease) → still virulent.
- Digested RNA (RNase) → still virulent.
- Digested DNA (DNase) → no transformation.
Conclusion: DNA is the transforming principle.
1.3 Hershey & Chase (1952) — Confirmed with Phage
- Bacteriophage T2 infects E. coli.
- Two batches of phages: one grown with ³²P (labels DNA), other with ³⁵S (labels protein).
- After infection and blender-shearing, they measured which label entered bacteria.
- Only ³²P went in — DNA (not protein) is injected.
- Definitive proof: DNA is the genetic material.
2. DNA Structure — Watson & Crick (1953)
Built on:
- Chargaff's rules (A = T, G = C, A + G = T + C).
- X-ray diffraction photos by Rosalind Franklin & Wilkins.
2.1 The Double Helix
- Two polynucleotide strands wound as a right-handed double helix.
- Strands are antiparallel — one runs 5'→3', the other 3'→5'.
- Sugar-phosphate backbone on the outside; bases on the inside, paired by H-bonds:
- Adenine (A) – Thymine (T) with 2 H-bonds.
- Guanine (G) – Cytosine (C) with 3 H-bonds.
- Dimensions: diameter 2 nm; each base-pair rise 0.34 nm; 10 bp per turn; pitch 3.4 nm.
- Grooves: major and minor — where regulatory proteins bind.
2.2 Chemistry
- Nucleotide = phosphate + pentose + nitrogen base.
- In DNA sugar = deoxyribose; bases = A, G (purines), C, T (pyrimidines).
- In RNA sugar = ribose; T → U (uracil).
3. DNA Packaging
- Human diploid genome ≈ 3.1 × 10⁹ bp; total length ~2.2 m stretched. Must be packed into a nucleus < 10 μm.
- Prokaryotes — no true nucleus; DNA (as nucleoid) is negatively charged and organised with positively charged proteins into supercoiled loops.
- Eukaryotes — DNA wraps around histones (H2A, H2B, H3, H4) as an octamer (2 of each). Each nucleosome has ~200 bp of DNA. H1 histone binds between nucleosomes.
- Nucleosome + linker DNA = a "beads on a string" appearance.
- Further coiling → 30-nm fibre → looped domains → chromatin → chromosomes.
- Euchromatin — loosely packed, lightly stained, transcriptionally active.
- Heterochromatin — densely packed, darkly stained, transcriptionally inactive.
4. DNA Replication
DNA must copy itself faithfully before every cell division. Watson-Crick model implied "semi-conservative" replication — each daughter gets one old + one new strand.
4.1 Meselson & Stahl (1958)
- Grew E. coli for many generations in ¹⁵N medium — all DNA became heavy.
- Shifted to ¹⁴N medium.
- After one generation: DNA was intermediate density (hybrid).
- After two generations: 50% intermediate + 50% light.
- Only semi-conservative replication fits — confirmed unambiguously.
4.2 Mechanism (in E. coli)
- Initiation at the origin of replication (ori).
- Helicase unwinds the double helix → replication fork.
- Topoisomerase (gyrase) relieves supercoiling ahead.
- Primase lays down short RNA primers.
- DNA polymerase III adds nucleotides only in the 5'→3' direction.
- Leading strand: continuous synthesis toward the fork.
- Lagging strand: synthesised discontinuously, in short Okazaki fragments away from the fork.
- DNA polymerase I replaces RNA primers with DNA.
- DNA ligase seals the nicks between Okazaki fragments.
Replication is amazingly accurate (~1 error in 10⁹ bases) thanks to polymerase proof-reading + mismatch repair.
5. Transcription (DNA → RNA)
5.1 Basics
- Only one strand of the DNA (the template strand, 3'→5') is transcribed.
- The other strand — the coding (sense) strand — has the same sequence as the RNA (except T → U).
- Catalysed by RNA polymerase, which synthesises RNA 5'→3'.
5.2 Prokaryotic vs Eukaryotic Transcription
Prokaryotes:
- Single RNA polymerase makes mRNA, rRNA, tRNA.
- No nucleus → transcription and translation are coupled (happen simultaneously).
- Sigma factor helps polymerase find the promoter.
- Rho protein helps terminate.
Eukaryotes:
- Three RNA polymerases:
- RNA pol I — rRNA (except 5S).
- RNA pol II — mRNA precursor (hnRNA).
- RNA pol III — tRNA, 5S rRNA, snRNAs.
- Primary transcript (hnRNA) has introns (non-coding) and exons (coding).
- Processing before export:
- 5' Capping — 7-methyl guanosine added.
- 3' Tailing (polyadenylation) — ~200 A residues.
- Splicing — spliceosome removes introns and joins exons.
- Mature mRNA is exported to the cytoplasm for translation.
6. The Genetic Code
Deciphered by Nirenberg, Khorana, Holley, and others in the 1960s.
Properties
- Triplet — 3 bases (codon) = 1 amino acid.
- Degenerate — most amino acids have >1 codon.
- Unambiguous — each codon codes for one specific amino acid.
- Universal — same code across species (rare exceptions in mitochondria/some protists).
- Non-overlapping and comma-less — read in fixed frame, no gaps.
- Initiation codon — AUG (also codes for methionine).
- Termination codons — UAA, UAG, UGA (do not code for any amino acid).
7. tRNA — the Adapter
Predicted by Crick, discovered later.
- 2D structure: cloverleaf — with an anticodon loop and an amino-acid attachment site (3'-CCA).
- 3D structure: L-shape.
- Each tRNA is charged by a specific aminoacyl-tRNA synthetase — pairing the correct amino acid to the correct anticodon.
8. Translation
Occurs on ribosomes. Ribosome = 2 subunits (60S + 40S in eukaryotes; 50S + 30S in prokaryotes).
8.1 Stages
- Initiation — small subunit binds mRNA at AUG; initiator tRNA (with Met/fMet) enters the P-site; large subunit joins.
- Elongation — next aminoacyl-tRNA enters A-site → peptide bond forms (catalysed by 23S rRNA — a ribozyme) → tRNA in P-site moves to E-site and leaves → ribosome shifts one codon (translocation).
- Termination — release factor recognises stop codon → polypeptide released.
The polypeptide then folds into its 3D shape (sometimes with help of chaperones) and may undergo post-translational modifications.
9. Regulation of Gene Expression
Cells don't need all proteins all the time. Regulation occurs at multiple levels — for our syllabus, focus on prokaryotic transcriptional control via the lac operon.
The Lac Operon (Jacob & Monod, 1961)
In E. coli, three structural genes are transcribed together as a single mRNA:
- z — β-galactosidase (hydrolyses lactose → glucose + galactose).
- y — permease (imports lactose).
- a — transacetylase.
Upstream:
- p — promoter (RNA pol binds here).
- o — operator (repressor binding site).
- i — regulator gene, transcribed independently, produces the repressor protein.
Absent lactose: repressor binds the operator → blocks polymerase → no transcription. Present lactose: some lactose enters the cell → converted to allolactose → binds the repressor → repressor releases the operator → transcription ON.
This is negative regulation with an inducer — a classic on/off switch.
10. Human Genome Project (1990–2003)
- Massive international effort to sequence the entire human genome.
- Sequenced by two consortia (public + Celera).
Key findings
- ~3.1 × 10⁹ base pairs in the haploid genome.
- ~30,000 genes — far fewer than the original estimate of 1 lakh.
- ~2% of DNA codes for proteins — most is regulatory, structural, or non-coding.
- Chromosome 1 has the most genes (~2,968); Y has the fewest (~231).
- SNPs (single nucleotide polymorphisms) — millions, useful for tracing ancestry and disease predisposition.
- Two randomly selected humans are 99.9% identical at DNA level.
11. DNA Fingerprinting
Alec Jeffreys (1985) developed the technique.
- Based on VNTRs (Variable Number Tandem Repeats) — short DNA sequences repeated a variable number of times among individuals.
- Steps:
- Isolate DNA from sample (blood, hair, semen, saliva).
- Digest with restriction enzymes.
- Separate fragments by gel electrophoresis.
- Transfer to nylon membrane (Southern blot).
- Hybridise with a labelled VNTR probe.
- Detect by autoradiography — the resulting band pattern is the "fingerprint".
Applications: forensic identification, paternity testing, tracing pedigrees, wildlife conservation, and evolution studies.
Key take-aways
- DNA is the genetic material — proved by Avery et al. and Hershey–Chase.
- Structure = function: the antiparallel double helix with A-T, G-C pairing enables fidelity, replication, and coding.
- Central dogma: DNA → RNA → Protein, with reverse transcription in retroviruses.
- Prokaryotes use one RNA polymerase; eukaryotes use three plus extensive mRNA processing.
- The genetic code is triplet, degenerate, and (nearly) universal.
- Regulation examples: the lac operon is a classic negative-inducible system.
- The Human Genome Project and DNA fingerprinting brought molecular biology into medicine, forensics, and everyday life.