Lecture 49: Transformation and Transduction Flashcards
Wednesday 19th February 2025
Who demonstrated the transforming principle?
Griffith
Describe Griffith’s experiment
- R cells (rough colonies) are non pathogenic
- S cells (smooth colonies) are pathogenic
- When mice were injected with R cells they were fine, because R cells are non-pathogenic
- When mice were injected with S cells they died, because S cells are pathogenic
- When mice were injected with heat-killed S cells, they were fine, because dead S cells are non-pathogenic.
- When mice were injected with R cells and heat killed S cells, the mice died
- This is because the DNA of the S cells was taken up by the R cells. This transformed the R cells into a pathogenic form
What happened after the Griffith experiments?
This experiment led to the discovery that DNA is the genetic material (later confirmed by Avery, MacLeod, and McCarty in 1944).
What do bacterial cells have to be in order to undergo transformation?
They have to be competent
What is transformation?
- a process in which bacteria take up and incorporate foreign genetic material (usually DNA) from their environment into their own genome.
- This allows them to gain new traits, such as antibiotic resistance or virulence.
- It is a form of horizontal gene transfer, meaning genetic material is passed between organisms without reproduction.
What is competence?
The ability of a bacterial cell to take up extracellular (‘naked’) DNA from the environment.
Give some examples of bacteria that are naturally competent
Bacillus subtilis
Streptococcus pneumoniae
Neisseria gonorrhoeae
Haemophilus influenzae
For bacteria that aren’t competent, what can happen?
Competence can be induced in bacteria that aren’t competent. For example, for biotech purposes with E. coli.
What is recombination ?
Recombination is the process by which genetic material is exchanged between DNA molecules, leading to new genetic combinations.
What is the advantage of recombination in bacteria?
Recombination result in more “fit” organisms and increases genetic diversity i.e. would serve similar purpose as eukaryotic recombination during meiosis
What are the benefits of bacteria recombination?
Drug resistance (bacteria can survive antibiotics).
Novel virulence characteristics (bacteria can become more infectious).
Novel metabolic capabilities (bacteria can break down new substances for survival).
What is horizontal gene transfer?
the process by which bacteria and other organisms transfer genetic material between each other, rather than passing it down from parent to offspring (which is vertical gene transfer). HGT is a major driver of bacterial evolution, allowing for rapid adaptation.
What are the types of horizontal/lateral gene transfer?
- Transformation
- Transduction
- Conjugation
Does horizontal gene transfer happen in eukaryotes?
It does happen, but it is rare
When do bacterial cells usually gain competence?
Usually, competence happens just before entry to stationary phase i.e. around the time they stop growing
This is because this is the point where their resources are running out
Describe differences in bacterial cell competence
Some e.g. Bacillus become competent for hours but only about 20% of the cells
Others e.g. Streptococcus all cells become competent but only for short amount of time
Most species not competent
Why do bacteria become competent at the entry to stationary phase?
- Because as the cells enter stationary phase, they know they are at a risk of dying.
- So they use quorum sensing, the ability to regulate genes based on ppopulaiton density.
How does B subtillis become competent?
- Cells secret ComX (small pheromone) so that its concentration increases in the environment
- As bacterial population density increases, ComX accumulates in the surroundings.
- Bacteria have ComP receptors and ComX will bind to these receptors.
- This will lead to a
reaction of events leading to changes in gene expression. - This change in gene expression will result in cells becoming competent and they can now undergo transformation, and take up foreign DNA from their environment.
(This system ensures that competence occurs only when bacterial population density is high, increasing the chances of successful gene transfer.)
Apart from ComX, what is another pheromone that can bind to b. subtilis?
CSF
What type of DNA is taken up for transformation?
- DNA is released during bacteria cell lysis
- Competent bacteria can bind and import these DNA fragments, potentially incorporating them into their genome through homologous recombination.
What type of DNA will b subtillis take up for transformation?
- B. subtilis will take up DNA of any origin
Why so some bacteria only take up DNA from their own species for transformation?
Because there is the risk of bacteria obtaining harmful phage DNA
Which bacteria will only take up DNA from their own species? (transformation)
Neisseria gonorrhoeae and Haemophilus influenzae will only take up from same species
How can the bacteria distinguish between DNA from another bacteria of the same species and from a different species?
Because it recognises specific DNA sequences. A short sequence of nucleotides is recognised by the receptor on bacteria.
Describe the general mechanism of transformation
DNA bind to surface proteins on the cell
Depending on species single or
double stranded DNA enters the cell
The DNA will then Bind to competence-specific protein whilst inside the cell. This prevents the DNA from getting broken down.
The DNA will then recombine with the DNA of the bacteria.
Is it true that recombination may convey new traits like lactose intolerance?
Yes
How can new traits be acquired?
Gene fragment usually big enough to contain several genes
Recombination may convey new traits
What are phages?
Viruses that kill bacteria
What is tranduction?
genetic exchange
in bacteria mediated by
bacteriophages
Describe phages
Phage genomes can be single or
double stranded DNA or RNA
Genome can be small – a few kbp or big – hundreds of kbp
Can be circular or linear
What are possibly the most abundant organisms on earth?
Phages
Do phages co-exist with hosts in all environments?
Yes
What are phages influenced by?
characteristics of host
population dynamics
long term evolution
Phage therapy timeline…
Are phages very specific?
Yes, they usually recognise specific species or strains (need to know bacteria first)
Can phage therapy be used as treatment for many diseases?
Yes, but more research on phages needs to be conducted
Do phages work well on biofilm?
Yes
Are phages safe?
Yes, but only a few studies on its safety, so more research needs to be done
Does T2 phage infect and kill e.coli?
Yes
What does the head of lambda phage contain?
Head contains double stranded linear genome
Size ~48 kbp
What is the tail of lambda phage important for?
Tail important for interacting with E. coli
How can lambda phage infect bacteria?
Infection can be either lytic or lysogenic
(not unique to 𝝺 but only applies to some phages)
Describe the lytic cycle (phages; transduction)
- The phage attaches to the bacteria and inserts its DNA into the bacteria
- The once-linear DNA circularises.
- The circular DNA now undergoes replication, transcription and translation so that new virions of phage are produced.
- The bacterial cell lyses and releases the virions of phage
- The released phages can then infect other bacterial cells, continuing the cycle.
What is the phage restriction system?
a bacterial defense mechanism against bacteriophage (viral) infection.
Describe the phage restriction system
Bacteria have a defense system called the restriction-modification system.
When a bacteriophage injects its DNA into the bacterial cell, the bacterium can cut the foreign DNA using restriction enzymes
These enzymes recognize specific DNA sequences and cleave phage DNA, preventing infection.
Host DNA protected because it is modified by methylation (many enzymes insensitive to methylation).
why is the phage restriction system important?
Restriction enzymes are now widely used in molecular biology for DNA cutting and genetic engineering.
The discovery of these enzymes helped develop recombinant DNA technology.
What is a lysogen?
strain of bacteria carrying a lysogenic phage e.g. E. coli (𝝺)
What is a prophage?
phage in lysogenic state
Describe the lysogenic cycle
- Phage attaches to bacteria and inserts its DNA
- The DNA becomes integrated into the host bacteria’s genome and new daughter cells will inherit this phage,
- No new virions are produced.
- E coli will live normally and healthily until something triggers the lytic cycle, but it is very rare and unlikely that something will trigger the lytic cycle.
When a bacteria has undergone the lysogenic cycle, how is the lytic cycle suppressed?
The lytic cycle is suppressed by expression of phage protein that stops lysis
This protein also suppresses lysis by other phage of the same type infecting the cell
This is perhaps because carrying a prophage is probably
an advantage to the host
What decided whether or not a bacteria goes through the lytic or lysogenic cycle?
Depends on genetic and environmental factors:
Nutritional state, UV light etc.
Is it true that most phages dont carry genes that are damaging to humans?
Yes, which is why phage therapy should have few side effects.
Is it true that only some bacteria can undergo transduction?
Yes
What are the 2 types of transduction?
Generalised transduction and specialised transduction
What is generalised transduction?
Transfer of any DNA to the recipient cell by lytic or lysogenic phage
What is Specialised transduction?
Transfer of specific genes via lysogenic phages
Describe generalised transduction
- Phage infects e coli cell.
- The phage DNA is replicated, and the host cell DNA is degraded.
- Virions are packaged, and occasional bots of host DNA are packed by mistake.
- The bacterial cell bursts (lyses), releasing the newly formed phage particles. Some of these phages contain bacterial DNA instead of viral DNA.
- The defective phage infects a new bacterial cell, but instead of injecting viral DNA, it injects the previous host’s bacterial DNA.
- If this DNA is incorporated into the new host’s genome via recombination, it can lead to genetic changes in the recipient cell.
What are some key points to remember about generalised transduction?
This process does not produce new phages because the transducing phages lack phage DNA.
The transferred bacterial DNA may carry new traits, like antibiotic resistance or virulence factors.
Generalized transduction is a random process, meaning any part of the bacterial genome can be transferred.
The probability of a specific genetic marker being transferred is very low (1 in a million to 1 in 100 million cells).
Describe specialised conduction
- specialized transduction only transfers genes near the phage’s integration site.
- A bacteriophage infects a bacterial cell
- Instead of immediately killing the host, the phage integrates its DNA into the bacterial genome at a specific site.
- The integrated phage DNA is called a prophage, and the bacterium continues to grow and divide, copying the phage DNA along with its own.
- Under certain conditions (e.g., stress, UV exposure), the prophage excises itself from the bacterial genome to enter the lytic cycle (where it produces new virus particles).
- However, sometimes the excision is imprecise, and the phage accidentally removes nearby bacterial genes along with its own DNA. At the same time, some of the phage’s own genes may be lost, making it defective.
- The defective phage DNA (which now includes some bacterial genes) is replicated and packaged into new virus particles. Since some essential viral genes are missing, these new phages cannot complete a full infection cycle.
- The defective phages are still capable of infecting another bacterium.
- They inject their DNA (which includes the bacterial genes from the previous host) into the new bacterial cell.
- The bacterial gene carried by the phage may integrate into the recipient’s genome through homologous recombination.
- If successful, the recipient bacterium gains new genetic traits, such as antibiotic resistance or the ability to metabolize new substances.
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Key messages
Horizontal gene transfer is a natural process that allows bacteria to diversity their genes and essential for our biotech exploitation
Genetic material taken up is inserted into the genome via recombination: Transformation, transduction and conjugation
Phages are viruses that prey on bacteria, but they can defend themselves
Phages are lytic or lysogenic (know the lifestyles!)
