Molecular Biology and Microbial Genetics: Study Guide

Unit 2.1 – Molecular Biology

Genotype vs Phenotype

The relationship between genotype and phenotype is fundamental in molecular biology. The genotype refers to the genetic makeup of an organism, while the phenotype encompasses observable traits resulting from gene expression.

• Genotype: The set of genes (DNA) present in an organism; encodes information for traits.

• Phenotype: The physical and biochemical characteristics expressed; determined by proteins produced from genes.

• Pathway: Genotype → proteins → phenotype.

• Example: A gene encoding an enzyme for pigment production results in colored cells (phenotype).

Nucleic Acid Terminology

Nucleic acids are polymers of nucleotides, which are the building blocks of DNA and RNA. Understanding their structure is essential for grasping genetic processes.

• Nucleotide: Composed of a phosphate group, a 5-carbon sugar (deoxyribose or ribose), and a nitrogenous base.

• Nucleic acids: DNA and RNA; polymers of nucleotides.

• Genes: Segments of DNA that code for proteins.

• Chromosomes: Long DNA molecules containing many genes.

• Genome: All genetic material in a cell.

DNA Structure

DNA is a double-stranded molecule with a unique structure that allows for accurate replication and information storage.

• Double stranded: Two complementary strands.

• Antiparallel: Strands run in opposite directions (5'→3' and 3'→5').

• Hydrogen bonds: Hold base pairs together (A–T, G–C).

• Sugar-phosphate backbone: Provides structural support.

• Double helix: Twisted ladder shape.

• Base pairing: Adenine (A) pairs with Thymine (T); Guanine (G) pairs with Cytosine (C).

DNA Replication

DNA replication is a semi-conservative process, ensuring genetic fidelity during cell division.

• Semi-conservative: Each new DNA molecule contains one old and one new strand.

• Leading strand: Synthesized continuously.

• Lagging strand: Synthesized discontinuously in Okazaki fragments.

Key Enzymes in DNA Replication

• Gyrase: Relieves twisting tension ahead of replication fork.

• Helicase: Unwinds DNA strands.

• Single-stranded binding proteins: Stabilize separated strands.

• Primase: Synthesizes RNA primer for DNA polymerase.

• DNA polymerase: Adds nucleotides (5'→3'), proofreads for errors.

• DNA ligase: Joins Okazaki fragments on lagging strand.

Accuracy of Replication

• Complementary base pairing ensures correct nucleotide incorporation.

• DNA polymerase proofreading and repair mechanisms correct errors.

DNA vs RNA

DNA and RNA are both nucleic acids but differ in structure and function.

• Similarities: Both are polymers of nucleotides; have sugar-phosphate backbone; carry genetic information.

• Differences:

  • DNA: Double-stranded, deoxyribose sugar, thymine (T).

  • RNA: Single-stranded, ribose sugar, uracil (U); leaves nucleus to participate in protein synthesis.

Gene Expression

Gene expression involves transcription and translation, converting genetic information into functional proteins.

• Transcription: DNA → mRNA.

• Translation: mRNA → protein.

• Central Dogma: DNA → RNA → Protein.

Transcription Steps

• Initiation: RNA polymerase binds to promoter region.

• Elongation: RNA strand is synthesized.

• Termination: RNA polymerase stops; mRNA is released.

RNA Polymerase vs DNA Polymerase

• Similarities: Both synthesize nucleic acids.

• Differences:

  • RNA polymerase synthesizes RNA; does not require a primer.

  • DNA polymerase synthesizes DNA; requires a primer.

Prokaryotic vs Eukaryotic Transcription

• Prokaryotic: Occurs in cytoplasm; no introns; operons allow one mRNA to encode multiple proteins.

• Eukaryotic: Occurs in nucleus; introns removed; one gene encodes one protein.

Introns & Exons

• Introns: Noncoding regions; removed during RNA processing.

• Exons: Coding regions; remain in mature mRNA.

Genetic Code

The genetic code is the set of rules by which information encoded in mRNA is translated into proteins.

• Codon: Sequence of three bases specifying one amino acid.

• Start codon: AUG (methionine).

• Stop codons: UAA, UAG, UGA.

• Redundancy: Multiple codons can code for the same amino acid.

Translation

Translation is the process by which ribosomes synthesize proteins using mRNA as a template.

• Initiation: Ribosome binds to start codon.

• Elongation: tRNA brings amino acids; peptide bonds form.

• Termination: Ribosome encounters stop codon; protein is released.

Simultaneous Transcription & Translation in Prokaryotes

• Occurs because prokaryotes lack a nucleus; both processes happen in cytoplasm.

Separation in Eukaryotes

• DNA is in nucleus; RNA must exit nucleus before translation.

Types of RNA

• mRNA (messenger RNA): Carries genetic code from DNA to ribosome.

• rRNA (ribosomal RNA): Structural and catalytic component of ribosomes.

• tRNA (transfer RNA): Brings amino acids to ribosome during translation.

Gene Regulation

Gene regulation allows cells to control which genes are expressed and when.

• Constitutive genes: Always expressed (ON).

• Regulated genes: Turned ON or OFF as needed.

Operon Concept

• Operon: Group of genes controlled together by a single regulatory region.

Inducible Operon (lac operon)

• Repressor is normally active (operon OFF).

• Inducer (lactose) inactivates repressor; operon turns ON.

• Example: Lactose metabolism genes.

Repressible Operon (trp operon)

• Repressor is normally inactive (operon ON).

• Corepressor (tryptophan) activates repressor; operon turns OFF.

• Example: Tryptophan synthesis genes.

Catabolite Repression (Lac Operon Conditions)

Catabolite repression regulates the lac operon based on glucose and lactose availability.

Glucose	Lactose	Lac Repressor	cAMP Levels	CAP	Lac Operon Expression
-	-	Active	High	Active	OFF
+	-	Active	Low	Inactive	OFF
+	+	Inactive	Low	Inactive	LOW expression
-	+	Inactive	High	Active	ON (maximum expression)

• Quick rule: Lactose turns the operon ON; glucose keeps it LOW; best expression occurs when lactose is present and glucose is absent.

Unit 2.2 – Microbial Genetics

Genomic Organization of Prokaryotes

Prokaryotes have a unique genomic structure, including chromosomal DNA, plasmids, and bacteriophages.

• Chromosomal DNA: Single, circular chromosome located in the nucleoid; contains thousands of genes.

• Plasmids: Small, circular, extrachromosomal DNA; replicate independently; often carry antibiotic resistance genes.

• Bacteriophage: Viruses that infect bacteria; may integrate into bacterial chromosome and transfer genes.

Genetic Change in Bacteria

Bacteria can undergo genetic changes through mutation and gene transfer, leading to new strains and species.

• Mutation: Permanent change in DNA sequence.

• Gene transfer: Exchange of genetic material between bacteria.

• Results in new strains, species, and antibiotic resistance.

Connection Between Genetic Change & Disease

• Genetic changes can increase virulence, produce toxins, confer antibiotic resistance, and allow immune evasion.

• Example: Mutation and gene transfer can create more dangerous pathogens.

Spread of Genetic Changes

• Vertical gene transfer: Parent to daughter cells during replication.

• Horizontal gene transfer: Between unrelated bacteria; rapid spread of traits; major cause of antibiotic resistance.

Mutation Terms

• Mutation: Permanent change in DNA sequence.

• Spontaneous mutation: Occurs naturally during replication.

• Induced mutation: Caused by external agents (mutagens).

• Mutagen: Chemical or physical agent causing mutation.

• Mutation rate: Frequency at which mutations occur.

Types of Mutation

• Missense: One base change; different amino acid.

• Nonsense: Base change creates stop codon; protein shortened.

• Silent: Base change but same amino acid; no protein change.

• Frameshift: Insertion or deletion shifts reading frame; usually severe.

Mutagens

• Base (nucleoside) analogs: Mimic normal bases; cause mispairing.

• Intercalating agents: Insert between DNA bases; cause frameshift mutations.

• Chemicals that alter bases: Change base structure; cause incorrect pairing.

• Radiation:

  • UV radiation: Causes thymine dimers; DNA distortion; mutations if unrepaired.

  • X-rays/Gamma rays: Break DNA strands; cause rearrangements.

Nucleotide Excision Repair

• Removes damaged DNA segments.

• Replaces with correct nucleotides.

• Example: Repair of thymine dimers caused by UV radiation.

Gene Transfer Mechanisms

• Transduction: Transfer of DNA by bacteriophage.

  • Generalized: Random bacterial DNA transferred.

  • Specialized: Specific genes transferred.

• Conjugation: Direct cell-to-cell contact; requires sex pilus and F plasmid; Hfr cells transfer chromosomal DNA.

• Transformation: Uptake of naked DNA from environment; requires competent recipient; DNA may integrate or remain as plasmid.

Plasmids – Important Features

• Circular DNA; independent replication.

• Carry accessory genes (e.g., antibiotic resistance).

• Transferable between cells.

R Plasmids (R Factors)

• Plasmids carrying antibiotic resistance genes.

• Clinical significance: Spread multidrug resistance; major healthcare problem.

Transposons (“Jumping Genes”)

• Structure: DNA segments flanked by insertion sequences; contain transposase gene.

• Function: Move within genome; move between plasmid and chromosome.

Relationship: Transposons + R Plasmids + Antibiotic Resistance

• Transposons often carry resistance genes.

• Can insert into plasmids.

• Plasmids transfer between bacteria.

• Rapid spread of resistance; major cause of superbugs.

Additional info: Horizontal gene transfer and mobile genetic elements (plasmids, transposons) are key drivers of bacterial evolution and the emergence of antibiotic-resistant pathogens.