Studied by 4 people
Semi-conservative replication
Keeps half of the original DNA, and half is newly synthesized DNA
What is the role of Helicase?
Enzyme that disrupts hydrogen bonds between complementary bases to separate the strands of the DNA molecule.
What is the role of DNA Polymerase?
Synthesizes complimentary daughter strands for each parent strand - adding 1 nucleotide at a time
What direction does DNA Polymerase move?
5’ to 3’
What is the leading strand?
The strand of DNA which DNA polymerase builds towards the replication fork continuously
What is the lagging strand?
The strand of DNA which DNA polymerase builds away from the replication fork, in segments
What are Okazaki fragments?
Short segments of DNA synthesized from the lagging strand
DNA Primase
Creates a primer (short chain of nucleotides) on the template strand and provides the site where DNA polymerase can bind to and start adding nucleotides.
DNA polymerase III
Binds to template strand on the primer and adds nucleotides to build daughter strand + proofreads after each nucleotide has been added to the chain
DNA Polymerase I
Replaces RNA nucleotides with DNA nucleotides on the lagging strand, proofreads newly synthesized DNA
DNA Ligase
Joins Okazaki fragments together
PCR
Polymerase Chain Reaction
3 stages of PCR
Melting, Annealing, Elongation
PCR Melting stage
Heating the DNA (to 95ºC) until hydrogen bonds break
What enzyme is used in PCR and why?
Taq polymerase is used because it is resistant to high temperatures (will not denature)
PCR Annealing stage
Cooling to 54ºC, allow primers to bind (prevents DNA from pairing up again)
PCR Elongation Stage
Heat up to 72ºC (optimum temperature for taq polymerase), taq polymerase replicates the DNA
Purpose of Gel Electrophoresis
To separate and analyze mixtures of DNA of different lengths
Gel Electrophoresis Procedure
Use of a tank with voltage (positive and negative on one side)
DNA is placed into wells near the negative side
DNA moves towards positive side (bc of slight negative charge from phosphate)
Small fragments travel further, bigger fragments are slower
