DNA replication Flashcards
What are the 3 possible models for DNA replication?
conservative (separate)
semi-conservative (mixed)
dispersive (one after another)
What was Meselson and Stahls question?
Is the mechanism of DNA replication conservative, semi-conservative, or dispersive
What did Matthew Meselson and Franklin Stahl do?
They used different isotopes of nitrogen to do their experiment looking into DNA replication
What method did Mathew Meselson and Franlin Stahl use?
1- Grow bacteria in media containing 15N: make “Heavy” DNA
2- Transfer them to media containing 14N: new DNA will be “light”
3- Separate heavy and light DNA molecules by ultracentrifugation
What’s the process of ultracentrifugation?
Using the force of central gravity (by spinning at certain speeds) to separate molecules in a solution based on their molecular weight (density)
*top of tube- less dense
What did Meselson and Stahl observe?
After 1st generation: sigle hybrid band discounted the conservative model
After 2nd generation: 2 separate band discounted the dispersive model
4th generation confirmed final conclusion
i.e. semi conservative replication is the one
What are the enzymes involved in DNA replication?
DNA polymerase
DNA ligase
Primase
Helicase
Topoisomerase
Singlr-strand binding protein
What does Primase do?
Primase synthesises short RNA sequences called primers on the single stranded DNA template.
then the primers act as a starting point for DNA polymerase (which can only add nucleotides to an existing strand)
what is the mechanism of DNA polymerase?
add nucleotides one at a time in 5’-3’ direction
Using template strand to form H-bonds: tells us which base to add next
tens or hundreds of nucleotides per second
*always add onto the 3’ of the new DNA strand
What does Ligase do?
it joins the loose ends together into a single strand of DNA (creating phosphodiester bonds between adjacent nucleotides)
What do Single stranded binding proteins do?
SSB binds to the separated strands, preventing them from reannealing.
What does Helicase do?
breaks hydrogen bonds between the two DNA strands, separating them.
What does Topoisomerase do?
relieves pressure from unwinding around the replication bubble by making and resealing breaks in the DNA
What are the leading strands?
5’-3’ DNA synthesis points towards the replication fork and can proceed continuously
what are the lagging strands?
5’-3’ SNA synthesis points away from the replication fork and must therefore be discontinuous (primed numerous times)
What are the Okazaki fragments?
the pieces of DNA that are stuck together to make up the Lagging strand of replication
Explain replication
1- helicase opens the template strand and breaks H bonds
2- single strand binding proteins prevent the strands from sticking together
3- Primase is adding primers
4- DNA polymerase is building the new strand, forming hydrogen bonds with the template strand
5- leading strands adds to replication fork continuously
6- lagging strand points away from the replication fork
7- primer is removed and gaps are filled in with ligase
what problem is caused to linear chromosomes during replication?
-Primer removal at the end of the chromosome leaves a gap that cant be filed in (there is no DNA polymerase coming along to fill in that piece
-So on every round of replication, a little piece is lost from the end of the chromosome (on leading strand- 5’)
-This is a problem for the lagging strand at each end of the linear DNA
*primase can only add to the 3’ ends
What are Telomeres? and what are they important for?
Short DNA sequences (6 nucleotides size) that are repeated over and over at the ends of the chromosomes
They serve as protective caps, preventing chromosome degradation and fusion.
What is telomerase?
its an enzyme that can lengthen telomeres
preventing them from shortening too much as a result of DNA replication - this allow these cells to divide indefinitely.
However in most body cells telomerase activity is limited, contributing to the aging process.
