Genetics Lecture 5: Chromosome Mapping in Prokaryotes and Eukaryotes

Genetic Analysis of Mapping in Bacteria and Bacteriophages

Enormous Diversity of Bacteria

Bacteria are among the most diverse and abundant organisms on Earth, occupying a wide range of habitats and playing critical roles in ecosystems and human health.

• Bacteria constitute one of the three major evolutionary lineages, alongside Archaea and Eukarya.

• In the average human, bacterial cells outnumber human cells by approximately 9:1.

• Most bacteria reside in the intestines, but the skin, mouth, and respiratory tract also host diverse bacterial communities.

• Bacteria are found in soil, aquatic environments, as parasites, symbionts, and even inside other life forms.

• Some bacteria cause diseases in plants and animals, while others are essential for nutrient cycling and environmental processes.

Additional info: Bacteria are prokaryotes, meaning they lack a nucleus and other membrane-bound organelles. They are always haploid, which allows mutations to be expressed directly.

Bacterial Genomes

Bacterial genomes are typically small, haploid, and circular, which facilitates genetic analysis and manipulation.

• Bacteria have diverse shapes, such as coccus (spherical), bacillus (rod-shaped), and spirillum (spiral).

• There are approximately 40 million bacterial cells in a gram of soil and a billion cells in a milliliter of fresh water.

• Estimated total bacterial cells on Earth: $6 \times 10^{30}$.

• Metagenomics is the collective analysis of genomic DNA from entire microbial communities, providing insights into diversity and function.

• The Human Microbiome Project has identified over 5,000 bacterial species in humans, with individual variation in species composition.

Example: Variations in the human microbiome have been correlated with disease, diet, and drug treatment. Some genome changes are associated with obesity.

Bacteria as Experimental Organisms

Bacteria and viruses are ideal for genetic studies due to their rapid reproduction, small genomes, and ease of manipulation.

Advantage	Description
Rapid reproduction	Allows quick generation of data
Many progeny	Facilitates statistical analysis
Haploid genome	Mutations are expressed directly
Asexual reproduction	Isolation of pure strains is simplified
Small genomes	Easy to sequence and manipulate
Medical importance	Relevant for disease studies
Genetic engineering	Production of commercial substances

Example: Escherichia coli is the most studied bacterium, commonly found in the intestines of warm-blooded animals. Lab strains are typically non-pathogenic and can grow in both aerobic and anaerobic conditions. E. coli is prototrophic, meaning it can grow in minimal media, and divides every 20 minutes.

Plasmids and Gene Transfer in Bacteria

Plasmids are small, circular DNA molecules capable of independent replication and often carry genes such as those for antibiotic resistance.

• Plasmids range from 0.05% to 10% of the size of the bacterial chromosome.

• Plasmids can be transferred between bacteria, facilitating the spread of traits like antibiotic resistance.

Additional info: Plasmids are key tools in genetic engineering and biotechnology.

Gene Transfer Mechanisms in Bacteria

Bacteria can exchange genetic material through several mechanisms, which are crucial for genetic mapping and analysis.

• Vertical gene transfer: Transmission of genetic material from parent to offspring during reproduction.

• Horizontal gene transfer: Introduction of genetic material from unrelated individuals or different species, including conjugation, transformation, and transduction.

Bacterial Culturing and Mutant Analysis

Bacterial strains are classified based on their nutritional requirements and ability to synthesize compounds.

• Prototrophs: Wild-type bacteria that can synthesize all compounds needed for growth from simple ingredients.

• Auxotrophs: Mutant strains lacking one or more enzymes required for metabolizing nutrients; require supplemented media for growth.

Genetic Recombination in Bacteria

Bacterial reproduction is typically asexual (binary fission), but genetic recombination can occur through several mechanisms.

• Conjugation: Direct transfer of DNA between two bacterial cells via a conjugation tube.

• Transformation: Uptake of free DNA fragments from the environment by competent cells.

• Transduction: Transfer of bacterial DNA by bacteriophages (viruses that infect bacteria).

Conjugation: F Factor and Cell Types

Conjugation involves the transfer of genetic material between donor and recipient cells, determined by the presence of the F (fertility) factor.

Factor	Characteristics	Role in Conjugation
F+	Present as separate circular plasmid	Donor
F-	Absent	Recipient
Hfr	Present, integrated into bacterial chromosome	High-frequency donor
F'	Present as plasmid carrying some bacterial genes	Donor

• Episome: A plasmid capable of integrating into the bacterial chromosome (e.g., F factor).

• During conjugation, the F factor is transferred from F+ to F- cells, converting recipients to donors.

• Hfr cells have the F factor integrated into their chromosome, leading to high-frequency recombination.

• F' cells contain plasmids with some bacterial genes, resulting from improper excision of the F factor.

Example: Conjugation between Hfr and F- cells can result in gene mapping by interrupted mating experiments.

Transposable Elements in Bacterial Genomes

Bacterial genomes contain transposable elements, which can disrupt gene function and rearrange genomes.

• Insertion sequences (IS): Simple transposable elements with inverted repeats and a transposase gene.

• Composite transposons: Two IS elements flanking a gene, often for antibiotic resistance.

• Transposons can cause deletions, inversions, and other genome rearrangements.

Gene Mapping by Interrupted Conjugation

Interrupted conjugation experiments allow mapping of bacterial genes based on the timing of gene transfer.

• Genes closer to the origin of transfer are transferred earlier during conjugation.

• By interrupting mating at specific times, the order and relative distances of genes can be determined.

Transformation and Gene Mapping

Transformation involves the uptake of extracellular DNA by competent cells, leading to genetic recombination.

• DNA fragments bind to receptors on the cell surface and enter the cell.

• One strand is degraded, and the other pairs with homologous sequences in the host genome.

• Recombination produces a heteroduplex DNA molecule.

• Gene mapping is possible by analyzing co-transformation frequencies, which are inversely proportional to the distance between genes.

Bacteriophages and Transduction

Bacteriophages (phages) are viruses that infect bacteria and can mediate gene transfer through transduction.

• Lytic cycle: Phage reproduces by lysing the host cell.

• Lysogenic cycle: Phage integrates into the host genome and replicates with it.

• Generalized transduction: Random fragments of bacterial DNA are packaged into phage particles and transferred to recipient cells.

• Transferred DNA can recombine with the recipient's genome, allowing gene mapping.

Example: Plaques on bacterial lawns indicate successful infection and can be used to estimate phage density.

Summary of Conjugation Outcomes

Mating Cells	Cell Types Present After Conjugation
Two F+ cells	F- cell becomes F+
One Hfr cell and one F- cell	No change (unless entire chromosome is transferred)
Two F' cells	F- cell becomes F'

Additional info: In Hfr x F- conjugation, the F- cell only becomes F+ if the entire chromosome is transferred, which is rare.