Genetic Recombination Flashcards
What happens when the 2 Holliday Junctions are resolved…
(1) The SAME way?
(Same point on both strands)
No crossover
Original molecules released
They have ALTERED regions -
A footprint of the exchange
What do DSBs initiate?
Recombination
Initiated by endonuclease which cleaves the recipient strand
Spo11 in meiosis
What does the exonuclease (working with helicase) do in recombination?
5’ end resection
(Nibbles away one strand on either side of the DSB)
Forming two ss tails with 3’ OH ends
One nibbled end invades a homologous region in the other (donor duplex)
How is heteroduplex DNA formed?
Exonucleases form TWO ss tails with 3’ OH ends
ONE nibbled end invades a homologous region in the other (donor) duplex
(Single-strand invasion!)
Heteroduplex formed - Base pairing between DIFFERENT parental strands
How is a Holliday Junction formed from Heteroduplex DNA?
When Single-Strand Invasion happens, one of the donor duplex strands is displaced : D LOOP
D loop is EXTENDED by DNA synthesis (replacing degraded material)
Once it can pair with the other side, that end is captured
The second strand of the gap is filled by more SYNTHESIS and ligation
Now there are two recombinant joints - HOLLIDAY JUNCTIONS
What happens when the 2 Holliday Junctions are resolved…
(1) In OPPOSITE ways?
(Strands nicked at recombinant joint)
A CROSSOVER is produced
What is Branch Migration?
The migration of a Recombinant Joint along the DNA Duplex
Which proteins recognise and resolve Holliday Junctions?
RuvA - Recognises & binds.
Its 2 tetramers sandwich the DNA.
RuvB - A helicase, catalyses branch migration
RuvC - Cleaves junctions
How does NHEJ repair DSBs?
Enzyme detail!
DEGRADING:
Ku binds the ends
Recruits DNA-PK (a kinase) and Artemis (a nuclease)
These degrade damaged DNA
HEALING:
Limited Microhomology lets ends come together (3-4bp)
Nucleases trim any extra
Polymerase fill any gaps
Ligase IV fills any gaps
What is the disadvantage of NHEJ?
Information can be lost from break site - Mutagenic
Biggest problem in GERM class and G1 cells without 2 copies of a chromosome yet
NHEJ is normally fine in somatic cells. But how can it cause issues?
Homologous recombination can occur between TRANSPOSABLE ELEMENT regions
What is Site-Specific Recombination?
Recombination between DEFINED sequences - mediate VERY SPECIFIC rearrangements
What THREE kinds of rearrangements can Site-Specific Recombination mediate?
INTEGRATION of a sequence
EXCISION of a sequence
INVERSION if the inserting DNA sequences sites are inverse of the genomic sequence
What enzymes catalyse Site-Specific Recombination?
Tyrosine + Serine RECOMBINASES
What is a real-world example of Site-Specific Recombination?
- Integration & Excision?
Phage LAMBDA Lysogeny
Integrates & Excises from bacterial genome
What is a real-world example of Site-Specific Recombination?
- Inversion
(a) Flagellar phase variation in Salmonella
Direction controls expression/suppression of different flagella types (express A, repress B and then the inverse) - Escape imm System
(b) Tail fiber variation in Phage Mu
Inversion in middle of S gene - flip between two types to infect more varied hosts
What happens during Chromosome Dimer Resolution in Bacteria?
XerCD recombinase
MEDIATES re-recombination between dif genes
This resolves chromosome dimers, which CAN’T segregate
What 4 levels of BACTERIAL Transposon complexity did we study?
- Encode TPase only
- Encode drug resistance, flanked by 2 transposase sequences (1 defective)
- Encode drug resistance, transposase, other genes like Resolvase (SS-Recombination system for insertion)
- A bacteriophage (uses TPosition)
( ! ) What FOUR mechanisms of recombinations did we study in this course?
- Homologous Recombination
- NHEJ
- Site-Specific Recombination
- Transposition