Important Diagrams: Molecular Basis of Inheritance

Table of Contents
1. Polynucleotide Chain
2. DNA Double Helix
3. Nucleosome
4. Hershey & Chase Experiment
5. Semi‐Conservative DNA Replication (Watson & Crick Model)
6. Meselson and Stahl's Experiment
7. Replicating Fork
8. Transcription unit
9. Transcription in Prokaryotes (Bacteria)
10. Transcription in Eukaryotes
11. Genetic Code
12. tRNA- the Adapter Molecule
13. Translation
14. Lac operon
15.  Human Genome Project
16. DNA Fingerprinting
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Polynucleotide Chain

Polynucleotide is a polymer composed of repeating units called nucleotides. Each nucleotide consists of three components: a five‐carbon sugar (deoxyribose in DNA, ribose in RNA), a phosphate group attached to the 5′ carbon of the sugar, and a nitrogenous base attached to the 1′ carbon. Nucleotides are linked by phosphodiester bonds between the 3′‐OH of one sugar and the 5′‐phosphate of the next sugar, creating a sugar‐phosphate backbone with directionality (5′ → 3′).

• Purines: adenine (A) and guanine (G); two‐ring structures.
• Pyrimidines: cytosine (C), thymine (T) in DNA, and uracil (U) in RNA; single‐ring structures.
• Sequence of bases along the backbone encodes genetic information.
• Polynucleotide chains have polarity: one end has a free 5′ phosphate and the other a free 3′ hydroxyl.

DNA Double Helix

The DNA double helix is the three‐dimensional structure formed by two antiparallel polynucleotide strands held together by hydrogen bonds between complementary bases. The model proposed by Watson & Crick (1953) explained base pairing and the helical geometry of DNA.

• Base pairing: A-T (two hydrogen bonds) and G-C (three hydrogen bonds).
• Strands are antiparallel: one runs 5′ → 3′ and the other 3′ → 5′.
• Right‐handed helix with approximately 10 base pairs per turn and a uniform diameter (~20 A).
• Major and minor grooves provide access for proteins that recognise specific base sequences.
• Chargaff's rules: in double‐stranded DNA total amount of A ≈ T and G ≈ C.

Nucleosome

Chromosomal DNA in eukaryotes is packaged into chromatin. The basic unit of chromatin is the nucleosome, which compacts DNA and regulates access to it.

• A nucleosome consists of ~146 base pairs of DNA wrapped ~1.7 turns around a histone octamer (two each of H2A, H2B, H3 and H4).
• Linker DNA between nucleosomes is associated with histone H1 and varies in length (~20-80 bp), producing the "beads‐on‐a‐string" appearance under low compaction.
• Further folding of nucleosomes forms higher‐order structures (30 nm fibre and beyond) leading to chromatin and chromosome formation.
• Post‐translational modifications of histone tails (acetylation, methylation, phosphorylation) influence chromatin structure and gene expression.

Hershey & Chase Experiment

The Hershey-Chase experiment (1952) provided strong evidence that DNA, not protein, is the hereditary material in bacteriophages.

• They used T2 bacteriophage labelled with radioactive isotopes: 32P to label DNA and 35S to label protein.
• Phages were allowed to infect E. coli, then the mixture was agitated in a blender and centrifuged to separate phage coats from bacterial cells.
• Radioactivity of infected bacteria was predominantly 32P, indicating DNA entered the cells and directed phage replication, while 35S remained with the phage coats.
• Conclusion: DNA is the material that transmits genetic information to progeny phages.

Semi‐Conservative DNA Replication (Watson & Crick Model)

Watson & Crick predicted that DNA replication would be semi‐conservative: each daughter DNA molecule would contain one original (parental) strand and one newly synthesised strand. Semi‐conservative replication preserves sequence information through complementary base pairing.

• Parental strands act as templates for the synthesis of new complementary strands.
• New strands are synthesised in the 5′ → 3′ direction by addition of nucleotides complementary to the template strand.
• Complementary base pairing (A-T and G-C) ensures faithful copying of information.

Meselson and Stahl's Experiment

The Meselson-Stahl experiment (1958) provided experimental proof for semi‐conservative replication using density gradient centrifugation.

• Escherichia coli were grown in medium containing heavy nitrogen (15N) so DNA became dense.
• Cells were shifted to medium with light nitrogen (14N) and allowed to replicate for successive generations.
• DNA samples were centrifuged in a CsCl density gradient; after one generation a single intermediate density band was observed (hybrid DNA), and after two generations both light and hybrid bands appeared.
• Results matched predictions of semi‐conservative replication and ruled out purely conservative or dispersive models.

Replicating Fork

The replication fork is the Y‐shaped region where parental DNA strands separate and new strands are synthesised. DNA replication involves a suite of enzymes and proteins that coordinate unwinding, priming, synthesis and ligation.

• Helicase unwinds the double helix at the fork.
• Single‐stranded binding proteins (SSBs) stabilise unwound DNA and prevent reannealing.
• Topoisomerase (gyrase) removes supercoils ahead of the fork by cutting and rejoining DNA.
• Primase synthesises short RNA primers complementary to the template to provide a 3′‐OH for DNA polymerase.
• DNA polymerase catalyses elongation by adding deoxynucleotides in the 5′ → 3′ direction. Leading strand is synthesised continuously; lagging strand is synthesised discontinuously as Okazaki fragments.
• DNA ligase joins Okazaki fragments by forming phosphodiester bonds.

Transcription unit

A transcription unit is a segment of DNA that is transcribed into an RNA molecule. It includes the promoter, the coding sequence (which becomes the RNA), and the terminator.

• Promoter: DNA sequence where RNA polymerase and associated factors bind to initiate transcription.
• Transcribed region: the portion of DNA used as template for RNA synthesis; includes exons and introns in eukaryotes.
• Terminator: signals where transcription ends.

Transcription in Prokaryotes (Bacteria)

In bacteria, transcription and translation are coupled. A single multisubunit RNA polymerase synthesises all classes of RNA; promoter recognition is mediated by the σ (sigma) factor.

• Core RNA polymerase + sigma factor = holoenzyme; sigma recognises conserved promoter sequences at -10 (Pribnow box) and -35 regions upstream of the transcription start site.
• Transcription proceeds in three stages: initiation, elongation, and termination.
• Termination occurs by ρ‐independent (intrinsic hairpin followed by U‐rich sequence) or ρ‐dependent mechanisms (ρ factor uses ATP to dissociate RNA‐DNA hybrid).
• mRNA in bacteria is often polycistronic (one mRNA encodes several proteins) and lacks extensive processing.

Transcription in Eukaryotes

Eukaryotic transcription is more complex and compartmentalised. Different RNA polymerases transcribe different types of genes and transcription requires multiple general transcription factors.

• RNA polymerase I: transcribes most rRNA genes (except 5S rRNA).
• RNA polymerase II: transcribes mRNA and many non‐coding RNAs; requires transcription factors (TFIID, TFIIA, TFIIB, etc.) and recognises promoters such as the TATA box.
• RNA polymerase III: transcribes 5S rRNA, tRNAs and other small RNAs.
• Primary transcripts (pre‐mRNAs) undergo processing: 5′ capping, 3′ polyadenylation (poly‐A tail), and splicing to remove introns by the spliceosome (snRNPs).
• Transcription is regulated by enhancers, silencers and chromatin structure; transcription factors and coactivators mediate controlled gene expression.

Genetic Code

The genetic code is the set of rules by which information encoded in nucleotide sequences (codons) is translated into amino acids. Key properties of the code are experimentally established and essential for translation.

• The code is based on triplets of nucleotides called codons; each codon specifies one amino acid or a stop signal.
• The code is degenerate: multiple codons can specify the same amino acid (redundancy), reducing the effect of some mutations.
• The code is generally non‐overlapping and read in a fixed reading frame beginning at a start codon.
• Start codon: AUG (codes for methionine and signals initiation of translation).
• Stop codons: UAA, UAG, UGA (do not code for amino acids; signal termination).
• The code is nearly universal with slight variations in mitochondria and some organisms.

tRNA- the Adapter Molecule

Transfer RNA (tRNA) functions as the adapter that interprets codons in mRNA and brings the corresponding amino acid to the ribosome during translation.

• tRNA has a characteristic cloverleaf secondary structure and an L‐shaped tertiary structure; key regions include the anticodon loop and the acceptor stem (3′ end) where the amino acid is attached.
• Aminoacyl‐tRNA synthetases charge tRNAs by covalently attaching the correct amino acid to the tRNA's 3′‐OH; this ensures fidelity in translation.
• Anticodon is a triplet of bases that pairs with the complementary codon on mRNA; wobble at the third position allows some tRNAs to recognise more than one codon.

Translation

Translation is the process by which ribosomes synthesise polypeptides using mRNA as the template. It proceeds in three major stages: initiation, elongation and termination.

• Ribosomes are complexes of rRNA and proteins: prokaryotic ribosomes are 70S (50S + 30S) and eukaryotic ribosomes are 80S (60S + 40S).
• Initiation: assembly of ribosomal subunits at the start codon (AUG) with initiator tRNA (initiator fMet‐tRNA in bacteria, Met‐tRNA in eukaryotes) and initiation factors.
• Elongation: charged tRNAs enter the A site, peptide bond formation transfers the growing chain to the tRNA at the A site (peptidyl transferase activity), and the ribosome translocates moving tRNAs through A → P → E sites.
• Termination: stop codon is recognised by release factors, polypeptide is released and ribosomal subunits dissociate.
• Polyribosomes (polysomes) are multiple ribosomes translating the same mRNA simultaneously, increasing protein synthesis efficiency.

Lac operon 

The lac operon in Escherichia coli is a classic example of gene regulation in prokaryotes. It is an inducible operon that controls utilisation of lactose as a carbon source.

• Components: structural genes lacZ (β‐galactosidase), lacY (lactose permease), lacA (transacetylase); promoter (P), operator (O), and regulatory gene lacI (repressor).
• In absence of lactose, lacI repressor binds to operator and blocks transcription of structural genes.
• When lactose (or its isomer allolactose) is present, it binds to the repressor causing an allosteric change and repressor releases operator; RNA polymerase transcribes the operon and enzymes for lactose uptake and metabolism are produced.
• Catabolite repression: presence of glucose reduces cAMP levels; low cAMP means CAP (catabolite activator protein) does not activate lac operon strongly, so glucose presence suppresses lac operon even if lactose is present.
• The lac operon illustrates negative and positive regulation and the integration of environmental signals into gene expression.

 Human Genome Project

The Human Genome Project (HGP) was an international effort to determine the complete sequence of the human genome and to identify all human genes. The project began in the late 20th century and achieved a near‐complete draft sequence in the early 2000s.

• Main goals: sequencing the 3.2 billion base pairs of human nuclear DNA, mapping genes to chromosomes, and developing technologies and databases for sharing genomic information.
• Key outcomes: a reference human genome sequence, improved sequencing technologies, identification of many genes and regulatory elements, and the foundation for genomics, personalised medicine and comparative genomics.
• Applications include identification of disease‐associated genes, improvements in diagnostics, pharmacogenomics and advances in biotechnology.

DNA Fingerprinting

DNA fingerprinting (also called DNA profiling) is the analysis of variable regions of the genome to identify individuals or relationships. It relies on genetic variation that is highly individual‐specific.

• Markers used: VNTRs (variable number tandem repeats), STRs (short tandem repeats), single nucleotide polymorphisms (SNPs) and RFLP patterns.
• Methods: earlier methods used restriction enzymes and Southern blotting (RFLP); modern methods commonly use PCR amplification of STR loci followed by capillary electrophoresis.
• Applications: forensic identification, paternity testing, missing persons identification, population genetics and biodiversity studies.
• Reliability depends on using multiple independent loci and proper statistical interpretation of match probabilities.

Optional brief summary: This chapter presents the molecular basis of inheritance: how DNA structure and packaging enable storage of genetic information, how DNA is faithfully replicated, how genes are transcribed and translated into proteins, and how gene regulation and modern genomic tools (operons, genome sequencing, DNA profiling) allow organisms to respond to the environment and enable advances in biology and medicine.