1 Replication of the Genetic Material Genetic material must be duplicated for transfer into daughter...

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Replication of the Genetic Material

Genetic material must be duplicated for transfer into

daughter cells.

Complementary double-stranded DNA

makes this possible

A

A

A

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Possible Mechanisms for DNA Replication

Which is correct?

semiconservative conservative dispersive

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Meselson - Stahl Experiment (1958)

Nitrogen isotopes

Density gradient centrifugation - CsCl

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Meselson - Stahl Experiment

Definitely not conservative

Results also consistent with dispersive

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Meselson - Stahl Experiment

Supports only semiconservative model

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Replication of DNA in Eukaryotic Chromosomes

How is DNA arranged and replicated?

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Replication of DNA in Eukaryotic Chromosomes

Taylor, Woods, and Hughes (1958)

CHO harlequinchromosomes

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DNA Polymerase Chain Elongation

Chain growth - 5’ to 3’ direction

3’ 5’ 5’ 3’

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DNA Polymerase Chain Elongation

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Replication Origin

Bidirectional Replication from Origin

Single origin in

Prokaryotes

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Replication Origins - Replicons

Multiple origins in Eukaryotes

Replicons

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Primer Requirement

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Initiation of DNA Replication - E. coli

Initiator protein DnaA binds oriC

denaturation

DNA helicases unwind DNA

DNA primase makes primers

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Replication Fork - Okazaki Fragments

Initial primers

Repriming

Discontinuous synthesis Lagging strand

Continuous synthesisLeading strand

5’

5’3’

3’

5’

3’

5’

3’

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Activities of DNA Polymerases

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Types of DNA Polymerases

E. Coli DNA polymerase I - single polypeptideDNA polymerase III - three polypeptide holoenzyme

Both capable of chain growth and 3’-5’ exonuclease activity Only pol I has 5’ - 3’ exonuclease activity

Eukaryotes

> 15 polymerases, 3 involved in replication of nuclear DNA

pol - with primase, creates RNA/DNA primerpol and pol - extend primers on leading/lagging strand

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Joining of Okazaki Fragments

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DNA Replication in E. coli (Fig. 3.6)

Initiation

Elongation

Primer Removal

Ligation

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DNA Replication in E. coli

Figure 3.8. Replisome Model. DNA pol III on lagging-strand templateis finishing synthesis of an Okazaki fragment.

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Telomere Problem

Due to:Linear chromosomes & Requirement for RNA primers

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Resolving the Telomere Problem

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Rolling Circle Replication

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Rolling Circle Replication in Phage