Understanding:
• The replication of DNA is semi-conservative and depends on complementary base pairing
DNA replication is a semi-conservative process, because when a new double-stranded DNA molecule is formed:
- One strand will be from the original template molecule
- One strand will be newly synthesised
Semi-Conservative DNA Molecule
This occurs because each nitrogenous base can only pair with its complementary partner
- Adenine [A] pairs with thymine [T]
- Cytosine
[C] pairs with guanine [G]
Consequently, when DNA is replicated by the combined action of helicase and DNA polymerase:
- Each new strand formed will be identical to the original strand separated from the template
- The two semi-conservative molecules formed will have an identical base sequence to the original molecule
Conservation of Sequence by Complementary Base Pairing
Skill:
• Analysis of Meselson and Stahl’s results to obtain support for the theory of semi-conservative replication of DNA
The theory that DNA replication was semi-conservative was confirmed by the Meselson-Stahl experiment in 1958
Prior to this experiment, three hypotheses had been proposed for the method of replication of DNA:
- Conservative Model – An entirely new molecule is synthesised from a DNA template [which remains unaltered]
- Semi-Conservative Model – Each new molecule consists of one newly synthesised strand and one template strand
- Dispersive Model – New molecules are made of segments of new and old DNA
Three Proposed Models of DNA Replication
Meselson and Stahl were able to experimentally test the validity of these three models using radioactive isotopes of nitrogen
- Nitrogen is a key component of DNA and can exist as a heavier 15N or a lighter 14N
DNA molecules were prepared using the heavier 15N and then induced to replicate in the presence of the lighter 14N
- DNA samples were then separated via centrifugation to determine the composition of DNA in the replicated molecules
The results after two divisions supported the semi-conservative model of DNA replication
- After one division, DNA molecules were found to contain a mix of 15N and 14N, disproving the conservative model
- After two divisions, some molecules of DNA were found to consist solely of 14N, disproving the dispersive model
Results of the Meselson-Stahl Experiment
- The first step in DNA replication is to ‘unzip’ the double helix structure of the DNA molecule.
- This is carried out by an enzyme called helicase which breaks the hydrogen bonds holding the complementary bases of DNA together [A with T, C with G].
- The separation of the two single strands of DNA creates a ‘Y’ shape called a replication ‘fork’. The two separated strands will act as templates for making the new strands of DNA.
- One of the strands is oriented in the 3’ to 5’ direction [towards the replication fork], this is the leading strand. The other strand is oriented in the 5’ to 3’ direction [away from the replication fork], this is the lagging strand. As a result of their different orientations, the two strands are replicated differently:
An illustration to show replication of the leading and lagging strands of DNA.
Image credit: Genome Research Limited
Leading Strand:
- A short piece of RNA called a primer [produced by an enzyme called primase] comes along and binds to the end of the leading strand. The primer acts as the starting point for DNA synthesis.
- DNA polymerase binds to the leading strand and then ‘walks’ along it, adding new complementary nucleotide bases [A, C, G and T] to the strand of DNA in the 5’ to 3’ direction.
- This sort of replication is called continuous.
Lagging strand:
- Numerous RNA primers are made by the primase enzyme and bind at various points along the lagging strand.
- Chunks of DNA, called Okazaki fragments, are then added to the lagging strand also in the 5’ to 3’ direction.
- This type of replication is called discontinuous as the Okazaki fragments will need to be joined up later.
- Once all of the bases are matched up [A with T, C with G], an enzyme called exonuclease strips away the primer[s]. The gaps where the primer[s] were are then filled by yet more complementary nucleotides.
- The new strand is proofread to make sure there are no mistakes in the new DNA sequence.
- Finally, an enzyme called DNA ligase seals up the sequence of DNA into two continuous double strands.
- The result of DNA replication is two DNA molecules consisting of one new and one old chain of nucleotides. This is why DNA replication is described as semi-conservative, half of the chain is part of the original DNA molecule, half is brand new.
- Following replication the new DNA automatically winds up into a double helix.
This page was last updated on 2021-07-21