Term
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Definition
| In asexually reproducing haploid populations, slightly deleterious mutations accumulate, as selection will only act on the worst affected individuals |
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Term
| Why does crossing over occur? |
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Definition
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Term
| How many chiasmata are needed|? |
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Definition
| Around one for every chromosome arm |
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Term
| What is the Holliday junction? |
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Definition
| A four way DNA junction formed by reciprocal exchange of single strands between two DNA duplexes |
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Term
| What is the Holliday model of recombination? |
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Definition
Homologous chromosomes align
Strands cut same positions by endonuclease
Cut strands exchanged between chromatids
Strands rejoined by ligase form Holliday junction
Holliday junction migrates to extend region of strand exchange forming heteroduplex DNA
Junction resolved by strand cleavage, seperating chromatids, resolution depends on orientation of cut |
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Term
| What are the two outcomes of the Holliday model? |
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Definition
North:South cut - > splice
West:East cut - > patch |
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Term
| What are the main issues with the Holliday model? |
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Definition
Nuclease needs to be incredibly precise to cut two strands at identical postions
No ab4:4 products (patch only on one chromosome) expected, but are seen in real life |
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Term
| What is the Meselson-Radding model of recombination? |
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Definition
single nick initiates (endonuclease)
Chain displaced
Strand invades homologous chromosome
Invaded part of chromosome removed
Ligation to Holliday junction
Resolution at this point leaves Ab4:4, OR
junction migrates
N:S, E:W resolution leads to splice or patch |
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Term
| What do both the Holliday and Meselson-Radding models ignore? |
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Definition
Ionising radiation increases frequency of recombination
Ionising radiation causes DSBs |
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Term
| What is the double strand break model? |
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Definition
DSB intiates
Exonuclease widens gap and leaves 3' ss tails
3' ss end invades homologue, forming D loop
3' end primes DNA synthesis, extending D loop
Other single strand engages displaced strand to form double holliday junction
Resolution can lead to splice, patch or Ab4:4 patch depending on resolution in same or opposite orientation |
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Term
| What is synthesis dependent strand annealing? |
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Definition
| part of DSB model, most non-crosovers dont need to form holliday junctions to patch |
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Term
| What are the stages of recombination? |
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Definition
Pre-synapsis - formation of DSBs and resection
Synapsis - DNA strand exchange and formation of joint molecules
Post-synapsis - branch migration, resolution and mismatch repair |
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Term
| What enzymes are involved in the pre-synapsis stage? |
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Definition
| Endonuclease to break DNA and Exonuclease to extend gap |
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Term
| What proteins are needed during synapsis? |
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Definition
Pairing protein to pair DNA
DNAP to extend 3' ends |
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Term
| What enzymes are needed during post-synapsis? |
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Definition
Helicase/endonuclease to resolve/move holliday junctions or D loops
Mismatch repair enzymes |
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Term
| What are the three approaches to identifying recombination genes |
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Definition
Genetics - identify and study mutants
Biochemistry - model DNA substrates of recombination
Informatics - parasite off of real scientists |
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Term
| What is the pilus encoded by? |
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Definition
F+ - plasmid forn
Hfr- integrated into chromosome |
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Term
| What is the oriV and oriT? |
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Definition
| F plasmids origin of replication and transfer respectively |
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Term
| What is an experiment to measure recombination in E.coli? |
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Definition
Two sets of E.coli, one F-, with steptomycin reisistance, that cant make proline, the other Hfr with Str susceptibility
Pro is near OriT, Str is on opposite side
Mix, and incubate at 37 degrees for 30-40 mins allowing mating pairs to form
Hfr transfers to F-
Interrupt mating
Spread an agar with no proline and streptomycin
Hfr and F parent cells cant grow, only Pro+, StrR can, No# colonies is a measure of recombination |
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Term
| What is a method of screening for recombination deficient mutants? |
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Definition
Mutagenise a F-, thr, leu, StrR strain and allow survivals to form colonies. Grow these on a regular array, then print on a avlevet pad. Transfer the pad to a thin lawn of Hfr cells that are StrS on minimal agar + streptomycin
Only recombinant colonies will grow, and the original mutants can be recovered from the master plate |
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Term
| How does RecABCD affect ends-out recombination occur in Hfr crosses? |
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Definition
RecBCD processes ends of transferred DNA and loads RecA
RecA promotes invasion |
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Term
| What do muations in sbcA do? |
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Definition
| Activate the quiescent recW gene encoding a dsDNA exo that suppresses the recBC phenotype? |
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Term
| What do sbcBC or sbcBD double mutations do? |
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Definition
sbcB inactivates a ss DNA exo
sbcC/D inactivates a mutiple exo
Suprresses recBC phenotype |
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Term
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Definition
| promotes assembly of recA filaments |
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Term
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Definition
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Term
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Definition
| RecQ helicase unwinds duplexed and tetraplexed DNA |
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Term
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Definition
| Form a complex that degrades long DNA plaindromes that form hairpin structures and process convergent replication forks |
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Term
| Im just gonna miss the page on ruvABC rec G recN and priA |
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Definition
| Because it appears to be mostly disconnected from the rest of the lecture and not very useful in an essay |
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Term
| What is the SOS response to DNA damage? |
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Definition
| Normally, LexA binds promoters of SOS genes, but it is inactivated in SOS reponse allowing transcription, also of recA which binds ssDNA exposed by damage and catalyses autocleaveage of LexA |
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Term
| What occurs in the RecBCD complex during pre-synapsis? |
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Definition
RecC recognises Chi and signals RecD to stop
RecD signals RecB to cut DNA
RecB cuts where it is and continues unwinding DNA, loading recA |
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Term
| What is the alternate method of pre-synaptic processing for recombination? with recQ and J |
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Definition
| RecQ unwinds DNA, RecJ exonuclease digests 5' strand to leave 3' ss tails |
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Term
| Why can recQ and recJ not make up for a mutant in the recBCD system? |
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Definition
| They can only facilitate efficient recA loading in recBC sbcBC strains lacking Exo 1 and SbcCD (I dont know what this means) |
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Term
| hat is the alternate method of pre-synaptic processing for recombination? with recE and T |
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Definition
| RecE exo degrades 5' of dsDNA, recT promotes DNA annealing and strand exchange, both are activated by sbcA, suppressors of recBC |
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Term
| Why is RecA important for strand exchange? |
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Definition
| It polymperises on the ssDNA and nucleates it randomly until it forms a stable RecA:DNA filament, which it builds in a 5-3 direction. This filament promotes pairing with dsDNA, aligning the ssDNA with the homologous dsDNA, and it generates a D loop as base pair switching displaces one of the duplex strands |
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Term
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Definition
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Term
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Definition
| RecA homologue for meiosis |
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Term
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Definition
| Eukaryotic RecA homologue |
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Term
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Definition
| Displaces SSB (Which binds ssDNA and competes with RecA) and allows RecA to bind - normally for repair |
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Term
| How are interstrand crosslinks repaired in E.voli? |
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Definition
Gap intially bound by SSB
RecA displaces SSB with help from RecFOR
RecA catalyses strand exchange
UvrABC does Excision repair
3' extended by DNAP
RUVABC resolves holliday junction |
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Term
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Definition
Tetramer in solution
Four arms of holliday junction dock into grooves on concave surface
Central acidic pin facilitates strand separation |
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Term
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Definition
| Binds to holliday junction, forms hexameric ring structures that function as helicases, unwind holliday junctions with help of RuvA, also need ATP and Mg2+ |
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Term
| How does RuvA help RuvB do its shit? |
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Definition
| Binds to holliday junction, RuvB binds to it, Ruv A has central pin to help seperate strands |
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Term
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Definition
| Binds to junction DNA and cleaves it to nicked duplex products, strand specific but dual strand cleavageq |
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Term
| How does RuvC help RuvAB? |
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Definition
| forms a resolvasome complex and scans for target sequences during branch migration |
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Term
| RecG helicase does something Im so tired can I stop doing these now |
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Definition
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Term
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Definition
Its another helicase that binds to and unwinds Holliday junctions because we need to know every single enzyme that does the same job as the previous enzyme we were just taught about.
It also unwinds D loops and molecule intermediates made by RecA, and R loops, where RNA invades dsDNA. |
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Term
| What can be used to avoid potentially dangerous recombination? |
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Definition
| PriA/C can restart replication |
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