Bacterial Genetics

Question 1

What is genome and in bacteria it is contained in what?

Answer 1

Genome is the totality of genetic information in an organism
CHROMOSOMAL
PLASMID

Question 2

What is the full definiton of a gene?

Answer 2

Gene
1. Unit of inheritance
2. Segment of the DNA/ RNA
3. Performs a specific function:
* Segment of the DNA Coding for an amino acid,
* Segment of the DNA coding for a peptide,
* Segment of the DNA a polypeptide /Others * Represented by sequence of nucleotide bases/ codon

Question 2

Chromosomes and DNA in bacteria

Answer 2

Coiled & super-coiled DNA,
Haploid in bacteria
Nil Recessive/ Dominant state

Bacteria DNA is closed & circular
DNA is coiled and super coiled to chromosome to make it fit into the cell

E.coli
4 million base pairs
1 mm long
over 1000 times larger that actual bacterial cell
DNA takes up around 10% of cell volume

Question 3

DNA replication

Answer 3

1. Chromosomal Replication starts with uncoiling of chromosome back into DNA by DNA helicase and gyrase
2. Uncoiled DNA is unzipped to create the replication fork
   DNA helicase-unzips the parental DNA strand that is used as a template

Leading stand synthesis is continuous (5’ to 3’)
Complimentary nucleotides are aligned to the DNA template
DNA polymerase joins aligned nucleotides to form growing DNA strand

Lagging strand synthesis is discontinuous (3’ to 5’)
RNA polymerase makes short RNA primer
DNA polymerase extends RNA primer by complimentary nucleotides alignment and linkage
DNA polymerase digests RNA primer and replaces it with DNA to form short DNA fragments called Okazaki fragments
DNA ligase seals Okazaki fragments

Question 4

What are Plasmids?

Answer 4

Plasmids are small, circular pieces of DNA that exist and replicate separately from the bacterial chromosome

Importance of Plasmid
Mutation (F-plasmid)
Resistance (R-plasmid)

Resistance or R-plasmids carry genes that confer resistance to certain antibiotics

R-plasmid usually has two types of genes:
R-determinant: resistance genes that code for
enzymes that inactivate certain drugs

RTF (Resistance Transfer Factor): genes for plasmid replication and conjugation.

Question 5

What are trasposons and examples

Answer 5

aka Transposable Genetic Elements or Jumping genes

Definition: they are pieces of DNA capable of moving from one location on the chromosome to another, from a plasmid to chromosome or vice versa and from one plasmid to another

Examples:
Insertion sequence
Episomes

Question 6

What are Insertion sequences (IS)?

Answer 6

Insertion sequences (IS) are jumping genes capable of attaching to other genetic elements of the cell from which they can move to another (transposition)

IS contains usually only one gene that codes for transposase, the enzyme that catalyses transposition.

The transposase gene is flanked by two DNA sequences called inverted repeats

Inverted repeats have nucleotide arranged upside-down and backward to each other.

Eg. Transposase helps in binding & excision of inverted repeats

Binding & excision of inverted repeats = MUTATION

Question 7

What are Episomes ?

Answer 7

• Episomes are fundamentally plasmids
• Are DNA segments
• Capable of independent existence and replication like plasmids
• Additionally, capable of integration into chromosomal DNA and excising from the chromosome
• Differs from IS in structure
• Like IS, can cause mutation

Question 8

Types of Mutation

Answer 8

1. Point mutations
2. Frameshift Mutations

Question 9

What are Point mutations?

Answer 9

1. Point mutations : aka base substitution
   occurs when a single nucleotide is replaced with a different nucleotide
   Results possibly in a mutant protein after transcription and translation.

Types of point mutations:
1. Silent Mutation:
* causes no change in the activity of the protein.
* Usually the result of a substitution occurring in the third location of the mRNA codon

2. Missence Mutation:
   * a nucleotide substitution that changes a codon so that it codes for a different amino acid in the protein.
   * usually results in a change of the activity of the protein
   * change may be harmful or beneficial
3. Nonsense Mutation:
   * same as a missense mutation except the resulting codon codes for a STOP signal
   * result in premature termination of translation
   * translated protein is shorter than usual and does not contain all the amino acids
   protein is most likely non-functional

Question 10

What are Frameshift Mutations?

Answer 10

Frameshift Mutations
caused by:
1. base insertion
2. base deletion

• An inserted or deleted nucleotide alters the triplet grouping of nucleotides into codons
• shifts the reading frame so that all nucleotides downstream from the mutation will be improperly grouped.
  *The result is a protein with extensive missense ending sooner or later in nonsense.

Question 11

Causes of mutation in bacteria

Answer 11

1. Errors made during replication, repair, or recombination
   Corrected by proof reading
   Even with proof reading, error abound
2. Exposures chemical agents (mutagens)
   Nitrous Acid:
   Nitrous Acid affects DNA complementation.
   The acid randomly modifies the base pairing
   adenine pairs with cytosine instead of thymine.
   A Base Analog:
   A base analog is a compound sufficiently similar to one of the four DNA bases but have different pairing properties.
   Examples:
   5-bromouracil is the analog of thymine but sometimes pairs with guanine
   2-aminopurine is the analog of adenine but sometimes pairs with cytosine.
   The incorporation of a base analog leads to a base pair substitution
3. Exposures to physical agents (Mutagen)
   UV Light:
   Exposure to direct UV light induces covalent linking between adjacent thymine nucleotides on a DNA strand forming a thymine dimer.
   These dimers cause the strand to buckle, disrupting normal base pairing.
   This prevents proper replication and transcription.
   Bacteria have enzymes to fix the damage created by UV light.
   An enzyme cuts the DNA at two point and removes the damaged portion.
   DNA polymerase synthesizes a new DNA segment using the healthly strand as a template.
   DNA ligase joins the new fragment to the old strand.
4. Genetic transfer in bacteria
   Genetic transfer in bacteria involves the mechanism by which DNA fragments are moved from a donor cell to a recipient cell
   Once donor DNA is inside the recipient, crossing over can occur (Recombination)

The result is a recombinant cell that has a genome different from either the donor or the recipient.

Question 12

What is the biological phenomenon associated with genetic element transfer in a bacterial population?

Answer 12

Transformation
Transduction
Conjugation

Question 13

What is Transformation?

Answer 13

The process by which a bacterium takes up naked DNA fragment from the environment

Sources of DNA in the environment:
bacterial cell death,
bacterial cell lyses,

Up taken DNA segment can align with homogenous section of recipient cell DNA

Aligned DNA can be incorporated/ integrated with recipient cell DNA (recombination)

Recombinant cell is a TRANSFORMED cell

TRANSFORMED cell is a MUTANT cell

Any up taken DNA that is not integrated into the recipient DNA will be degraded.

Recombinant cell /Transformed cell / Mutant cell has a different genetic makeup compared to Donor & Recipient.

All of the descendants of the recombinant cell will be identical to parent cell.

Recombination gives room to genetic diversity in bacterial population.

Question 14

What is a BACTERIOPHAGE?

Answer 14

Viruses capable of infecting bacterial cells/prokaryotes

Question 15

How can you classify BACTERIOPHAGES?

Answer 15

Lytic and lysogenic phages

Question 16

Describe the lytic cycle of a BACTERIOPHAGE

Answer 16

1. Bacteriophages bind to a host cells and injects it’s genetic material (DNA or RNA) into the cell
2. the phage DNA/RNA takes over the cell’s protein synthesis and replication machinery, forcing it to express the phage genes, produce phage proteins, and replicate the phage genetic material.
3. The newly produced phage proteins and DNA assemble into phage particles, after which the infected cell bursts, or lyses, releasing 100–200 new viral particles ready to infect other cells.
4. The cell lysis and release of progeny phage is called the lytic cycle of phage multiplication.

The lytic cycle is also known as the reproductive cycle of the bacteriophage

Question 17

Describe the lysogenic cycle of a BACTERIOPHAGE

Answer 17

1. The lysogenic cycle does not result in immediate lysing of the host cell.
2. Those phages able to undergo lysogeny are known as temperate phages. Their viral genetic material will integrate with host DNA and replicate along with it fairly harmlessly, or may even become established as a plasmid.
3. The virus remains dormant until host conditions deteriorate, perhaps due to depletion of nutrients; then, the endogenous phages (known as prophages) become active.

If the bacterial cell attempts to heal itself (curing), it will attempt to excise the viral genetic material by restriction endonuclease

5. At this point they initiate the lytic cycle, resulting in lysis of the host cell. As the lysogenic cycle allows the host cell to continue to survive and reproduce, the virus is reproduced in all of the cell’s offspring.

An example of a bacteriophage known to follow the lysogenic cycle and the lytic cycle is the phage lambda of E. coli.

Question 18

Enumerate the differences between the LYTIC & lysogenic cycles of BACTERIOPHAGES

Answer 18

Lytic Cycle:
1. Virulent phages
2. Phage infects host bacterium
3. Phage DNA replicates
4. Host cell’s machinery hijacked for phage production
5. New phage particles assembled
6. Host cell lyses (bursts), releasing phages
7. Phages infect nearby bacteria

Lysogenic Cycle:
1. Temperate phages
2. Phage infects host bacterium
3. Phage DNA integrates into host genome (prophage)
4. Prophage remains dormant
5. Host cell survives and continues to grow
6. Prophage replicates with host DNA
7. Can be induced to enter lytic cycle (e.g., UV radiation)

Question 19

What is Transduction?

Answer 19

Involves the transfer of DNA from one bacterium to another through a bacteriophage (phage)

A phage is a virus that infects bacteria.
Examples of Phage infecting E. coli

1. T4 phage
   -replicate by the lytic cycle
   - Cell death
2. Lambda phage
   -replicate by the lysogenic cycle
   -latent infection in the host until it breaks out in a lytic cycle

Question 20

What are the Types of Transduction?

Answer 20

1. Generalized
   - uses lytic cycle
   - any part of bacterial DNA can be transferred by transduction
2. Specialized
   - uses lysogenic cycle
   - Specific part of bacterial DNA can be transferred by transduction

Question 21

What is Conjugation?

Answer 21

• Transfer of genetic element between bacterial cells through conjugation bridge
  Donor & Recipient cell are of opposite polarity (+ & -)
  Conjugation bridge is fromed through sex pilus (Cf: Common pilus)

Conjugation bridge = Cytoplasmic bridge
Cytoplasmic bridge is temporary, dissolves after conjugation

Cell+ is the donor
Cell- is the recipient and becomes +

Donated genetic element is a plasmid/ episome

Not all bacteria can undergo conjugation
Presence of a special plasmid called the F plasmid is important for conjugation

Bacteria that have a F plasmid are referred to as F+ or male.
Those that do not have F plasmid are F- or female.

Conjugation event occurs when the male cell extends his sex pili and attaches to the female.

a temporary cytoplasmic bridge is formed through which the F plasmid is transferred from the male to the female.

When transfer is complete, the result is two male cells.

The F plasmid can behave as an episome.
When the F+ plasmid is integrated with the bacterial chromosome, the cell is called an Hfr cell (high frequency of recombination cell).

The F plasmid always insert at the same spot for a bacterial species.

The Hfr cell still behaves as a F+ cell, transferring F genes to a F-cell, but now it can take some of the bacterial chromosome with it.