Bacteriophage Lambda Flashcards

1
Q

What are bacteriophage Lambda?

A

Bacterial viruses that infect E.coli.

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2
Q

How can the phage be classified?

A

By various physical and molecular characteristics, but mainly classified by their appearance.

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3
Q

Outline the different appearances that phage can be classified through.

A

Mainly by their appearance:
- ICOSAHEDRAL HEAD, WITH A TAIL: Limited amount of nucleic acid you can fit inside the icosahedral head, so there are limitations to the duplication of DNA, due to the limited physical size of the head that the DNA has to be packaged into.
It is limited in the way it can evolve based on their structure, due to the physical size of the head, the DNA needs to be packaged into, so once the limit is reached, it cannot duplicate anymore.
This is the shape of the lambda phage.

-FILAMENTOUS PHAGE: I.e. long piece of nucleic acid, coated in proteins bound to the nucleic acid.
These are not limited in evolution because they can just get longer, they do not reach a limit on the amount of genetic material.

However they can also be classified by the type of nucleic acid they contain e.g. RNA/ DNA.

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4
Q

What are the two life cycles the phage can undergo?

A

Lytic and lysogenic.

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5
Q

How do both the lytic and lysogenic life cycles start?

A

They both need to start with getting binding to the maltoporin receptor on the outer membrane of the E.coli cell with its tail, and injecting the DNA into the E.coli.

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6
Q

Why can’t the filamentous phage inject its DNA into the E.coli as the icosahedral ones do?

A

Because they haven’t got the inbuilt system to dock on the membrane receptor, so they need to first unpackage the nucleic acid, then are taken in to the bacteria by unknown mechanisms.

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7
Q

How do we know that filamentous phage first unpackage their nucleic acid before transporting it into the bacteria?

A

Because in experiments, can see that for a brief period of time, the nucleic acids become susceptible to nucleases outside the cell, then are taken into the cell by an unknown mechanism.

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8
Q

Outline the differences between the lytic and lysogenic life cycle.

A

So they both start the same way, with injection of the phage DNA into the E.coli through the maltoporin receptor on the outer membrane.

The lytic: The phage genome is translated into the different viral components, allowing new virus particles to be synthesised, which will then form pores in the cell wall, allowing the release of the viral particles, hence killing the cell.

The lysogenic: The phage genome will be integrated into the host genome, and is replicated as the cell replicates, so is transferred to all the daughter cells, then when the cells are stressed etc. the lambda phage genome is excised from the host’s genome, and it essentially then follows the lytic cycle for the second half, resulting in cell lysis.

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9
Q

Why is the lytic cycle often called the reproductive phase?

A

Because in this phase, you are actively reproducing many different viral particles.

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10
Q

What is a virulent phage?

A

A phage that will always undergo the lytic cycle.

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11
Q

What is a temperate phage?

A

These have a decision as to which pathway to follow depending on the conditions of the host cell.
Most bacteriophage are temperate.

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12
Q

When could a temperate phage become a virulent phage?

A

If the temperate phage has a mutation.

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13
Q

What is a lysogen?

A

A bacterial cell containing the phage genome.

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14
Q

What is a prophage?

A

The piece of DNA that has been assimilated into the bacterial chromosome. It has the ability to become a phage again, but in its integrated state it is a prophage.

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15
Q

What is immunity in the context of lambda phage?

A

Lysogeny gives immunity to the E.coli from further infection from other bacteriophage particles, because once the prophage has been integrated into the genome, it protects it from further infection (by the production of a protein that stops another lytic cycle from occurring, and so it won’t go down another lytic cycle.

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16
Q

What is induction?

A

An event causing a lysogen to release the prophage, so that it becomes a separate viral genome in the bacterial cell.
A lysis event will generally occur after a period of lysogeny when the correct conditions are met.

17
Q

What decision must be made when infection occurs?

A

Whether to go down the lytic or lysogenic cycle.

18
Q

What determines whether the lytic or lysogenic cycle occurs?

A

In most cases, it depends on the conditions of the host cell.

19
Q

Why would it be advantageous for the phage to go down a lytic cycle?

A

Because hundreds of viral particles are produced in the bacterial cell, which are then released into the environment.

20
Q

In which cases would lysogeny be favoured?

A

When there are low levels of nutrients.

Also if there is a high multiplicity of infection (MOI).

21
Q

Why is lysogeny favoured when there are low levels of nutrients?

A

Because in these conditions, the bacteria don’t grow very quickly and so can go into a dormant phase where they are not making any proteins etc. and so no transcription occurs… these are things that the bacteriophage needs to make more viral particles.

So if the phage is injected into bacterial cells that are about to undergo dormancy, then the bacterial cell is essentially breaking down all components associated with an active life cycle, so before dormancy occurs, the phage genome can be integrated into the host chromosomes, then protected.

22
Q

Why does a high multiplicity of infection favour lysogeny?

A

Because the virus replicates at a much faster rate that the bacterial cells, and so it reaches a point of high MOI, whereby there are more viral particles present thatn there are bacterial cells, and so there is no point to undergo lysis at this point as there are not enough bacterial cells to infect for new viral particles.
So, if there is a high concnetration of bacteriophage and a low number of bacterial cells, then the viral particles are better off to be integrated into the bacterial genome, and so it would protect it from further infection from any other viruses, so the bacterial cells will continue to grow, and the daughter cells will also contain the prophage, and they would all be protected due to immunity.

23
Q

What happens if the bacterial population rises following a period of high multiplicity of infection?

A

The phages can be induced to undergo the lytic cycle.

24
Q

What are the two ways a plaque of the bacteria may appear like?

A

Clear or turbid.

25
Q

How are bacteria referred to as a group?

A

Colonies.

26
Q

What happens when you spread bacteria over a plate?

A

You will get a bacterial colony for each bacterial cell initially spread over the plate.

27
Q

What are bacteriophage cultures referred to as?

A

Plaques.

28
Q

How do you investigate bacteriophage cultures?

A

By spreading lots of bacteria on a plate, forming a solid surface of bacteria, then adding the phage particles to this.
Each time a phage infects the bacterial cells, you will get a hole in the bacterial wall… i.e. so it is the opposite to colonies… you are looking for holes.

29
Q

What kind of plaques are produced by virulent phage?

A

Clear plaque- because they will just keep killing the bacteria that are present in that point on the plate.

30
Q

What kindof plaques are produced by temperate phages?

A

They produce turbid plaques.
In the middle of this zone, is where there is the highest concentration of phage particles, where they kill off the bacterial cells.
It is likely that at one point, one of the bacterial cells will be converted to a lysogen, gaining immunity, and will hence grow in the centre of this clear plaque, so forming a central turbid plaque in the centre of the clear ones.

So, there is less bacterial growth than you have in the whole lawn, but now completely clear plaque due to the overgrowth of the lysogenic cells that have occurred.

31
Q

Describe the size and shape of the bacteriophage lambda genome?

A

Linear dsDNA, 48, 490bp.

32
Q

Where in the bacteriophage is the genome found?

A

Encased in an icosahedral head, with a tail particle attached.

33
Q

What happens when the genome is injected into the host cytoplasm?

A

It circularises, using the 12bp stranded regions at both the 3’ and 5’ ends, which anneal, so it is now 48,502 bp.

34
Q

Compare the configuration of the lambda phage genome in the phage particles in the host cell.

A

In the phage particle, it is linear, with a 12bp single stranded regions at both 5’ and 3’ ends, at 48, 490 base pairs.

In the host cytoplasm, the genome circularises, with hydrogen bonds forming between the two overhangs of 12bp, to form a circular genome of 48,502 bp.

35
Q

How many genes are expressed on the lambda phage genome?

A

46 genes.

36
Q

Where is the cos region found?

A

Between the lysis gene and the start of the head operon.

37
Q

Why must the genome be circularised?

A

Because if not circualties, then you won’t get expression of the later genes, as it requires expression of the promoter.

38
Q

What genes are found in the late genes?

A

This is a mega-operon that contains all the genes for all the head and tail proteins, i.e. phage particle production.

39
Q

What is the function of the control region?

A

To make decisions between the lytic and lysogenic life cycle.