Bacterial Genetics 3

Question 1

HGT / LGT

Answer 1

• horizontal / lateral gene transfer

- any process by which an organism transfers genetic material to another cell that is not its immediate offspring

Question 2

Bacteriophage

Definition

Answer 2

-a virus that infects bacteria, often shortened to phage

Question 3

Promiscuous

Definition

Answer 3

-has a broad host range

Question 4

Transduction

Definition

Answer 4

• mechanism by which chromosomal DNA can be transferred between bacteria, providing an opportunity for recombination
• a sexual process in bacteria mediated by bacteriophages

Question 5

Virus

Answer 5

• microscopic particle that can infect the cells of a biological organism
• cannot reproduce on their own, can only replicate by infecting a host cell
• consist of genetic material/nucleic acid contained within a protective protein coat, a capsid
• genetic material can be double stranded or single stranded and DNA or RNA

Question 6

Virus - MS2

Answer 6

• only has one capsid protein

- 3.6 k nts of RNA in genome

Question 7

What are the components of a T4 phage?

Answer 7

• head
• neck/collar
• core
• sheath
• end plate
• fibres

Question 8

What are the two types of infection by phages?

Answer 8

• virulent e.g. T2 & T4

- temperate e.g. lambda

Question 9

Portals

Answer 9

• surface molecules that act as receptors for bacteriophages

- do not exist only as portals for phage entry, they also serve some other important function

Question 10

Phage λ Receptor

Answer 10

-member of the sugar porin family that facilitates diffusion of mlltose

Question 11

Phage T4 Receptor

Answer 11

• OmpA

- a porin that provides a pathway for diffusion of small hydrophilic compounds in and out of cells

Question 12

Virulent Phage Infection

Answer 12

• leads inevitably to cell death
  1) adsorption of phage to host cell
  2) entry of phage nucleic acid into host cell
  3) phage proteins synthesised and phage genetic material replicated using host cell machinery and degradation of host cell genome
  4) assembly of progeny phage particles within the host cell
  5) lysis of host cell releasing progeny phage particles

Question 13

Phage Therapy

Definition

Answer 13

-use of lytic phages to treat pathogenic bacterial infections as a potential alternative to antibiotics

Question 14

Phage Therapy

Description

Answer 14

• the idea is to use a phage to infect and kill specific bacteria whilst not interacting with surrounding cells in the infected organism or harmless bacteria
• phages replicate quickly so only a small dose would be sufficient
• phages have a particular host range so would only b capable of infecting certain cells
• usage of antibiotics suggests that it would be possible for target cells to become resistant to phages
• but the difference between phages and antibiotics is that phages are able to evolve at a fast rate as well so could overcome some of the problems with antbiotic resistance

Question 15

MRSA

Answer 15

• about 1/3 of people are carriers
• infection occurs when MRSA gets into body tissues or the bloodstream
• individuals with a weakened immune system are the most at risk
• infection are often treatable with antibiotics (but not standard ones as MRSA is resistant to these)

Question 16

Transducing Phage

Answer 16

• in case of lysis by some phages (NOT T4) bacterial DNA from the degraded host genome can become accidentally packaged into the capsid with the phage genome
• this is rare and only happens occasionally
• P1 and P22 are examples of generalised chromosome transducing phages as they can carry any part of the genome
• P1 and P22 are often used in the lab to move chromosomal DNA (e.g. antibiotic resistance genes) between different strains

Question 17

Generalised Transduction

Answer 17

1) adsorption of phage to surface of donor bacterium
2) insertion of phage DNA into donor bacterium
3) degradation of donor bacterium genome, synthesis of phage proteins and replication of phage genome
4) formation of phage particles, in some case fragments of bacterial genome are packaged in the place of phage DNA
5) lysis of cell and release of progeny phage particles
6) phage particle containing bacterial DNA binds to surface of recipient bacterium
7) bacterial DNA from donor inserted into recipient bacterium
8) recipient bacterium is now a partial diploid, and there is sequence homology between the circular recipient genome and the linear donor strand
9) crossing over occurs at each end of the linear molecule and the donor strand is incorporated into the recipient genome by homologous recombination
10) the section of DNA that is swapped out for the donor strand is lost
11) the product of this process is a bacterium which contains DNA from two parent cells, the recipient and the donor

Question 18

Coliphages

Definition

Answer 18

-phages that infect E.coli

Question 19

Prophage

Definition

Answer 19

-name given to the temperate phage genome once it has been incorporated into the hosts chromosome

Question 20

Lysogenic Bacterium / Lysogen

Definition

Answer 20

-bacterium containing a prophage and growing normally

Question 21

Lysogeny

Definition

Answer 21

-the act of blocking the lytic cycle (via the synthesis of a repressor molecule) and integrating the phage genome into the bacterial chromosome

Question 22

Temperate Infections

Answer 22

1) phage binds to host cell and inserts DNA
2) the lytic response can begin now, or it can be supressed and the lysogenic response begins instead
Lysogenic Response
-integration of phage chromosome into the host cell genome
-cell under goes normal division
-until the lytic response is triggered
-the phage genome is excised from the bacterial chromosome and the phage enters the lytic cycle
Lytic Response
-replication of phage genome and breakdown of bacterial DNA
-synthesis of phage proteins
-assembly of progeny phage particles
-lysis of cell and release of progeny phages

Question 23

Production of Lysogen

Answer 23

i. e. how the prophage is formed
- there are regions of homology between the phage chromosome and the bacterial chromosome
- these regions of perfect homology are very small, only 7-15bp, too small for efficient homologous recombination
- so dedicated recombinase enzymes are needed that recognise the sequences of homology
- crossing over occurs at two points allowing the phage genome to be integrated into the bacterial chromosome

Question 24

Homologous Recombination

Answer 24

• nicking of parental DNA at the same place on each chromosome
• the same section on each chromosome is able to unravel
• each strand base pairs with the complimentary sequence on the opposite chromosome
• ligation at the nicking point
• the position of the chromosomes relative to each other changes such that is possible to make a single cut to separate the two recombinant chromosomes

Question 25

Production of Initial Lysate

Answer 25

i.e. excision of the phage chromosome from the bacterial chromosome upon initiation of the lytic cycle
i) Normal Out Looping
-the chromosome loops back on itself bringing the two -ends of the phage chromosome together
crossing over occurs
-the complete phage chromosome is removed exactly as it was before integration
ii) Rare Abnormal Out Looping
-as there are many IS elements in the genomes, it is possible for looping out to occur at different points to the integration
-so crossing over occurs at different places and the loop of DNA excised is not the same as the DNA that was integrated
-part of the phage genome may be left behind in the bacterial chromosome and part of the bacterial chromosome may be excised into the phage genome

Question 26

Specialised Transducing Phage

Definition

Answer 26

• a transducing phage that can only carry regions from the right or left hand side of the point of integration normally at a specific site
  e. g. lambda