DNA Replication and Repair1C part 1 Flashcards
What were the three models of DNA replication that researchers thought of?
- Semiconservative Replication
- Conservative Replication
- Dispersive Replication
Semi-conservative Replication+ configration
Each daughter strand remains paired with its complimentary parental strand.
one strand daughter, one strand parent
Conservative
after replication, both daughter strands pair up parent strand stay together
Dispersion
Daughter strand will have a mixture of parental and newly synthesized DNA
Meselsin Stahl Experiment tell me the procedure (3)
- Took normal coli cell DNA and made them divide in heavy nitrogen (N15) so all the DNA were heavy.
- DNA switched to light nitrogen and dna made after the switch would have to be made up of N14 as this would have been the only nitrogen available for DNA synthesis.
- measured the density of the DNA (and, indirectly, N14/N15) using density gradient centrifugation.
generation 0 meselson-Stahl (3)
- contents
-Band
-time
- All heavy N15 E coli DNA
- before the switch to light nitrogen
-produced a single band after centrifugation.
Generation 1 Meselson Stahl (3)
content
band
fit/reject
- 100% hybrid of light and heavy nitrogen
-fits with dispersive and semi conservative but not conservative
-single band middle
Generation 2 Meselson Stahl
- Two bands, 1 middle and 1 light band
- Know that it was conservative as in dispersive replication, all the molecules should have bits of old and new DNA, making it impossible to get a “purely light” molecule.
- 2 hybrid, 2 light
what do you expect over Generation 3/4 of the Meselson Stahl experiment?
over the third and fourth generations, we’d expect the hybrid band to become progressively fainter (because it would represent a smaller fraction of the total DNA) and the light band to become progressively stronger (because it would represent a larger fraction).
Why is the buffer important in the Meselson and Stahl experiment and what is it? (2)
Caesium Chloride solution forms a density gradient when centrifuged, the Cs ions will form an increasing concentration going down the tube and the DNA molecules will float to the density zones that match their own density. This allows separating similar molecules with small differences in density.
Since proteins are heavier they sediment in the heaviest, bottom-most layer while RNA being lighter is present at the topmost layer with the lowest density
spacing btwn the possible bands are
equal
Nucleotides can only be added to the new strand at the____ and thus DNA is
3’-OH end and thus, Dna is synthesized in a 5’-3’ direction
What end of the daughter cell is growing?
3’ end of daughter strand is growing
What provides energy for the formation of new phosphodiester bond
hydrolysis of pyrophosphate provides energy for the formation of new phosphodiester bond
During strand elongation, two phosphates are cleaved from the incoming nucleotide triphosphate and the resulting nucleotide monophosphate is added to the DNA strand.
pyrophosphate (1)
is just two phosphates (POP)
DNPT (4)
Name
type
what
dna
Deoxyribose nucleoside Triphosphate
DCTP
DGTP
DATP
DTTP
contains deoxyribose with 3 phosphate groups
lose 2 phosphate during DNA replication incorporation
DNA polymerases (5)
synthesizes daughter in 5’-3’
can only add bases on the 3’ OH end
reads template in a 3’-5’ end
cannot synthesize from scratch, it required a RNA primer that has a 3-OH group end untouched for DNA polymerases to bind and replicate.
Is an enzyme so it has an active site. Active site can catalyze 4 different reactions with the different DNTP. It binds to whatever the complimentary base is as that is the most energetically favoured.
Replisome (9)
- helicase
- primase
- single stranded binding protein
- DNA topoisomerase/Gyrase
- DNA Pol 3
- DNA Pol 1
- Sliding Camp
- Ligase
- molecular machines of enzymes that replicate DNA
Helicase
Unwinds the Double Helix by breaking hydrogen bonds btwn the 2 parental strands
Primase (2)
Synthesizes RNA primers for DNA polymerase
-provides the hydroxyl group
Single Stranded binding proteins
stabilizes single stranded DNA before replication (as single stranded can break) by preventing reannealing so that the strands can serve as template
DNA Topoisomerase/gyrase+ how it does this
removes supercoils that form ahead of the replication fork, relives torque of mainly circular DNA
does this by either making single stranded cuts or cutting both double strands and rehealing it.
DNA polymerase 3
synthesizes DNA by adding nucleotides to the new DNA strand
DNA polymerase 1
removes RNA primer and fills in the gap between with DNA
Sliding clamp (2)
attaches DNA pol 3 to DNA template to prevent it from dissociating. Without it, DNA poly 3 would synthesize a short segment before falling off
pushes DNA polymerase 3 to go faster and increase replication speed
Ligase+ present in… (2)
Joins the ends of DNA segments that DNA pol 1 synthesized after removing the RNA primer by forming phosphodiester bonds.
present in leading strand and in lagging strand.
How are the two strands different in DNA Replication?
one strand is synthesized continuously (leading)
one strand is synthesized discontinuously (lagging) in small DNA fragments
Why is there a lagging strand?
Because DNA polymerase can only synthesize in 5’-3’ direction
DNA replication steps (7)
- DNA helices unwinds double helix
- RNA primate lay down RNA primer
- Topoisomerase prevents dna twisting ahead of replication fork during unwinding
- dna poly 3 extends rna primer
- dna helicase continue to unwind resulting in leading and lagging strands synthesis by dna pol 3
- dna pol 1 removes rna primer of ozakaki fragments and fills in gaps with dNTPs
- DNA ligase seals nicks by reforming the phophodiester bond
