Conjugation, Transformation, and Transduction Flashcards
Conjugation
The transfer of genentic material between bacterial cells by direct cell to cell contactvia a sex pilus
- The F plasmid sex pilus conects with a specific site on the female recipient bacterial cell
- The sex pilus redracts through depolymerization forming a congugal pore in which the donor and recipient are in close proximity
- One strand of double stranded plasmid DNA is nicked by the enzyme nickase which binds specificly to th the origin of transfer
- The nicked piece of DNA is carried by nickase to the base of the conjugal pore
- DNa is passed into the recipient through the secretory apparatus in an energy depentent manner - ATPase activity
- Replication occurs
F Plasmid
The fertility factor that contains the conjugation machinery
The cell that contains the F plasmidd is the donor and produces the sex pilus
Rolling Circle Replication
Transfer of chromosomes through conjugation
- Recombination of the host genome must occur using a conjugative plasmid that has the following
- OriR – Origin of replication
- Tra – transfer machinery: all the genes that are required to make the sex pilus, nickase (replication machinery)
- If the plasmid integrates into the chromosome, then all of the transfer machinery is now located in the plasmid
- What facilitates the recombination into a chromosome?
- Insertion Sequence (a transposable element)
- An insertion sequence is a small piece of DNA (~1kb) that encodes a gene for an recombinase enzyme, transposase
- Also needs to have the inverted repeat that the transposase can bind to and recognize – and transpose the DNA
- If a chromosome has the same insertion sequence as a plasmid the two can recombine to make a larger circle which now has the transfer machinery
- The chromosome can now produce the sex pilus, and nickase which nicks the DNA and transfers the chromosome through the secretory pore in an ATPase dependent manner – the hybrid chromosome is transferred into the recipient
- If the two cells are kept in contact long enough, the entire chromosome can be passed to the recipient and can recombine
How to determine the relationship of genes
Interupted maiting
Select for the recipient (kill the donor)
At time points – Interupted maiting by vortexing
Can see patern of transfer
If vortex before 5 minutes will only see gene A transferred
After 5 minutes will see gene A+B transferred
Can set up order – mapping through interrupted maiting
Conjugation DNA
Never in the environment
Can be specific or general
Transformation
The genetic alteration of a cell resulting from the direct uptake and incorperation of exogenous DNA in the environment
DNA is naked
Can be specific or general
- Cell lysis occurs and DNA is released into the environment
- Bacterial cells become competent and the environmental donor DNA attaches to specific cell surface receptors
- DNA is taken into the cell as single stranded DNA during translocation across the inner membrane
- Recombination occurs - for homologous recombination to occur the incoming DNA contains regions of sequence similarity to the host genome. Recombination can also occur through additive integration - circular plasmids
Why is DNA in the environment?
- Cell lysis - programed or due to cell death
- Secretion - by T4S machinery into the environment
Stability of DNA depends on the environment
How do Bacteria take advantage of DNA in the environment?
- Food source – DNA can be used as a food source for the cell
- Biofilm formation – part of the matrix is made of DNA which is stabilized by histone like proteins, provides structural support (biofilms can be killed by nucleases)
- Allows for evolution – take up the DNA and can alter genetic material for genetic diversity
Competence
Competence is the ability of a cell to take up extracellular (“naked”) DNA from its environment.
The recipient becomess competent in expressing receptors for DNA
This is signaled by quarum sesnsing
The cells want large numbers of bacteria in the environment to become competent
Quorum sensing signals enter the cell and activate the genes for DNA receptors and genes that encode the machinery for DNA intake (only single stranded DNA gets into the cell), and machinery to stabilize the single stranded DNA
Cells that take up DNA
- Cells that will only take up their own DNA – species specific
- Cells that are non discriminate about the DNA that they will take up – whores (generally the DNA taken up is used as nutrients)
Transduction
The process by which DNA is transfered from one bacterium to another by a bacteriophage
Does not require physical contact between the cell dontating the DNA and the cell recieving the DNA
DNase resistant
Specific
- A bacteriaphage binds specifically to a receptor on the surface of the donor and injects the phage DNA into the donor
- The DNA canenter the lytic or lysogenic cycle
- If entering the lytic cycle, the viral genome replicates and produces progeny phages which induce lysis
- During the process of replication the host chromosome is sheered and the host cell chromosome will be packaged into phages (if it is the right size)
- Infectious particles are released and can go on to infect other donor cells
- Phage particles containing transducing particle are taken in by a recipient cell but cannont replicate - but recombination can occur
Adsorption
The adhesion of viral particals to a specific recepto on the surface of a bacterial cell
Infectious particle
Phage particles that contain the genome of the donor phage
infectious, and can infect the next host
Transducing Particle
Phage particle that have the genome of the host cell chromosomes are called transducing particles and cannot replicate when injected into the host cell
However recombination CAN occur
