Lesson 3: DNA Replication Flashcards
DNA in Prokaryotes
- Contain a single large double stranded DNA molecule in protein held in the cell’s nucleoid region (pseudo-nucleus)
- Contains one or more small circular DNA molecules called plasmids which float in the cytoplasm
DNA in eukaryotic cells
-Contains a double stranded DNA formed in the nucleus of the cells
-DNA binds with the protein Histone to form chromosomes during cell division
The possible Mechanism of DNA replication
1. Semi-conservative
The two strands of DNA separate and each serves as a template for the synthesis of a new complementary strand
The possible Mechanism of DNA replication:
2. Conservative
The entire molecule serves as a template for a new molecule
The possible Mechanism of DNA replication:
3. Dispersive
Two strands break into units that are replicated and reassembled with the new molecule alternating segments
Meselson and Stahl (1958)
Meselson and Stahl discovered the replication method by suing different isotope of N and E. Coli
-E. Coli was first grown in a medium containing “heavy” nitrogen (15^N). When extracted this DNA will be near the bottom of the solution of CsCl
-E. Coli extracted from E. Coli grown on normal N (14^N) will be on the less dense top of the solution of CsCl
—> the bacteria can controls the N content
-after time on the “heavy nitrogen medium”, the E. Coli were transferred to a normal N medium for one generation. Their DNA will contain 15^N and 14^N
-when placed in a CsCl solution there was one line halfway between the location of the 14^N and the 15^N location
—> eliminating the conservation model theory
Meselson and Stahl (1958) pt 2
-When grown for a second generation in 14^N medium and placed in a CsCl tube, two bands were observed.
-> one at the 14^N/15^N position and one at the 14^N position.
-this observation eliminated the dispersive model theory
The confirmed Mechanism of DNA replication
Semi-conservative model.
DNA replication in prokaryotes
-mostly plasmids which are circular molecules that can float around the cytoplasm
—> the replication process of these circular moles is referred to as the rolling wheel method
Rolling wheel method 1
-grp of initiation enzymes called topoisomerases recognize a 100-200 base nucleotide sequence called the replication origin
—> the enzymes bind to the DNA at this site and separates the 2 strands creating a replication bubble
Rolling wheel method 2: Elongation
DNA POLYMERASE III enters the bubble and binds to e/ parental Strand of DNA, they use each strand as a template in order to create a new complementary DNA strand one nucleotide at a time
Rolling wheel method 2: Elongation how the DNA POLYMERASE is replicated
5’ to 3’ direction, therefore the replication forks move away from e/o
Rolling wheel method Stage 3: termination
Once the replication forms meet the process is over and the two molecules separate
DNA replication in eukaryotes
-eukaryotic DNA is much larger than the prokaryotic DNA
-Long, large, double helical strands
-more complicated process
DNA replication in eukaryotes 1: Initiation
-DNA GYRASE (a topoisomerase) recognized an origin of replication sequence on the parent DNA (euk DNA has many origins of replication)
-a section of DNA unravels creating a replication bubble
-Helicase enters the replication and uncoils and separates the double helix
-Single stranded binding proteins (SSBP’s) bind to the separated DNA strands to prevent them from reattaching
DNA replication in eukaryotes 2: Elongation
-DNA Primase binds to e/ parent strand of DNA and creates an RNA PRIMER which is used as a starting point for replication
-DNA Polymerase III binds to both parental DNA strands and will create 2 new strands from the existing parental DNA templates
—DNA Polymerase III can only form a new strand in the 5’ to 3’ direction
DNA replication in eukaryotes Elongation: replication of the leading strand
-DNA polymerase III creates a new complementary DNA strand one nucleotide at a time
-this process is continuous and fairly fast since the replication fork is moving in the same direction as the DNA polymerase
DNA replication in eukaryotes: Elongation : Lagging strand pt 1
-for this Strand the DNA polymerase is moving in the opposite direction of the replication
-lagging strand is replicated in pieces called okazaki fragments
How does it know there’s a mistake in the DNA replication
Complimentary base pairing, and uses this to fix it
DNA replication in eukaryotes stage 3: Termination
Once the replication is complete the strands recoil into a double helix, however before that DNA polymerase III must reach an area of the DNA called telomere
—> the lifespan of the cell is dependent on the lifespan of the telomere regions because every time DNA replicates the new daughter strand is a few nucleotides shorter because the og RNA primer on the leading strand and the last RNA primer on the lagging strand are not converted to DNA
Telomere
Safe termination regions containing repeating TTAGGG sequences
DNA replication in eukaryotes 2: Elongation Problem
Parental strands are anti-parallel, meaning that the replication process is different for each strand.
3’-5’ parental strand = leading strand
5’-3’ parental strand = lagging strand
DNA replication in eukaryotes: Elongation : Lagging strand pt 2: DNA Primase
creates an RNA primer which is used as a starting point for the first DNA fragment
DNA replication in eukaryotes: Elongation : Lagging strand pt 3: DNA polymerase III
binds to the strand and forms a new strand from the starting point
— as more of the double helix is unwound, another RNA primer is formed by DNA Primase and another DNA polymerase createes another fragment
DNA replication in eukaryotes: Elongation : Lagging strand pt 4: DNA polymerase I
will then convert the RNA primers into DNA
DNA replication in eukaryotes: Elongation : Lagging strand pt 5: DNA ligase
pieces the okazaki fragments together
DNA replication in eukaryotes: Elongation : Lagging strand pt 6: DNA polymerase III returns
will then proofread e/ nucleotide and correct any mistakes in replication
