Lecture 12 - Genetic Recombination Flashcards
Why do genes need to have stability?
Genome conservation, DNA repair, void cancer and genetic disorders
Why is genetic diversity needed?
Meiosis, antibody diversity, adaptation and evolution
What are the 5 mechanisms of genetic recombination?
Homologous recombination, non-homologous end joining, transposition, site-specific recombination and independent assortment of chromosomes.
Homologous recombination - what happens after a double strand break?
The ends are processed by a nucleuses - the RecBCD complex.
The broken DNA can then invade an intact homologous DNA
Homologous recombination - what helps strand invasion?
Recombinant (RecA)
Homologous recombination - what happens after invasion?
The broken DNA is repaired by using the intact DNA as a template for replication and faithfully synthesise missing DNA.
Homologous recombination - what are Holliday junctions?
The 4 DNA branch structures
Homologous recombination - what can cleave Holliday junctions?
Resolvable called RuvABC
Homologous recombination - where can ruvABC cleave?
It has the choice between side ways and above/below - this will make different molecules
Homologous recombination - what happens if both Holliday junctions are cleaved on the same strand?
No-crossover - REMEMBER CHANGE STRANDS AT THE ARROWS
Homologous recombination - what happens if the RuvABC cuts different strands?
Crossover - REMEMBER YOU FOLLOW THE ARROWS
When does Homologous recombination take place?
DS break, ds DNA fragment, DNA single stranded gap
Non-Homologous end Joining - what does this repair
double stranded break and is good for diversity
Non-Homologous end Joining - what happens after a DS break?
The end are protected by Ku which keeps the ends next to each other.
Non-Homologous end Joining - what happens after Ku has bonded?
Sometimes they will naturally ligaments together, other times they need to be processed first
Non-Homologous end Joining - what happens during end processing?
This is carried out by different enzymes - nucleotides are added due to polymerase and removed by nucleuses. Nucleotides can be modified by phosphotase. This changes the original DNA
Non-Homologous end Joining - what happens after end processing?
Lig4 ligands the ends together.
What is transposition?
Movement of transposable elements within and between elements.
This is an origin of genetic diversity
site-specific recombination - what does it need at specific target sites to work?
Recombinase
site-specific recombination - what does recombinase do?
Binds at specific target sites and cleaves DS DNA at specific sequences which allows the change of order before they rejoin once more
site-specific recombination - what does the nature of the change rely on?
The orientation of the target sites - head to head or head to tail
site-specific recombination - what does a target site orientation of head to head mean?
They are going in the opposite orientation
site-specific recombination - what does head to tail orientation mean?
They are in the same orientation
site-specific recombination - what are the consequences of inversion (head to head) recombination?
There is an inversion of the sequence between the two target sites
Part of the target sites are also exchanged (arrows)
site-specific recombination - what are the consequences of deletion/ integrate (head to tail) recombination?
Line up the arrows - whether this is a deletion or an integration depends on the molecule.
The recombinase lines up the target sits creating a loop. This then leads to 2 products - a circle containing the DNA sequence between the 2 target sites (and one Target site) being excised and the original molecule without the DNA which previously sat between the target sites and only one target site.
site-specific recombination - can the 2 products recombine to form the original strand?
Yes
site-specific recombination in nature - e.coli replicate on the inside of the circle which means straight DNA can be bad for them. What template is used for the repair?
The inside one
site-specific recombination in nature e.coli - what is the product of this repair?
Non-crossover or crossover - these are still circular
site-specific recombination in nature - e.coli - what happens if there is crossover?
They can’t go into their own cell and therefore must be unlinked before that.
site-specific recombination in nature - e.coli - how do they unlink crossed DNA?
Site specific recombination - a specific head to head target site called dif and the recombination is done by xerCD recombinase which causes a second crossover which then lets them unlink.
Following infection of bacteria wha can bacteriophage lambda do?
Use its host to replicate in the lambic cycle or insert its DNA and force the host to replicate that.
How does the lambda bacteriophage integrate or excised from the bacteria it is infecting?
Site specific recombination
Lambda genomes recircularise after entering the bacteria genome. This now makes them able to enter e.coli chromosomes - how?
They line up their attp site with attB sites on the e.coli chromosomes and uses the int protein to insert themselves.
Is lambda and e.coli genome recombination reversible?
Yes
What are the composite sites formed called?
AttL and attR
Are attp and attB sites different?
Yes which means more enzymes need to be used for integration
Site specific recombination in flagellar phase variation in Salmonella? - where are the target sites?
Head to head
Site specific recombination in flagellar phase variation in Salmonella? - what does the fljb gene encode?
Protein which makes flagellum
Site specific recombination in flagellar phase variation in Salmonella? What does fljA encodes?
Regulator protein which represses fljc
Site specific recombination in flagellar phase variation in Salmonella? - what does fljc encode?
Another flagellum protein
Site specific recombination in flagellar phase variation in Salmonella? Where is the promoter of fljb?
In between 2 head to head target sites
Site specific recombination in flagellar phase variation in Salmonella? - what happens if the promoter is in front of fljb?
The flagellum is made up of fljb protein
Site specific recombination in flagellar phase variation in Salmonella? - what happens when the signal is inverted?
Fljb and fljA proteins aren’t produced and therefore fljc is no longer repressed and we can form fljc flagellum’s.
Bacteriophage Mu tail fibre variation - what type of recombination event - head to head or head to tail?
Head to head
Bacteriophage Mu tail fibre variation - this is made up of many protein - what codes for the constant part?
sc
Bacteriophage Mu tail fibre variation - this is made up of lots of proteins - what codes for the variable part?
so or sv+
Bacteriophage Mu tail fibre variation - what is the second protein?
U or U+
Bacteriophage Mu tail fibre variation - what does gin seen on the end of the tail encode?
Recombinase which can interrupt the genome between the target sites to encode either scSvU or ScSv+U+
