1 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

2 Possible Mechanisms for DNA Replication Which is correct? semiconservative conservative dispersive

3 Meselson - Stahl Experiment (1958) Nitrogen isotopes Density gradient centrifugation - CsCl

4 Meselson - Stahl Experiment Definitely not conservative Results also consistent with dispersive

5 Meselson - Stahl Experiment Supports only semiconservative model

6 Replication of DNA in Eukaryotic Chromosomes How is DNA arranged and replicated?

7 Replication of DNA in Eukaryotic Chromosomes Taylor, Woods, and Hughes (1958) CHO harlequin chromosomes

8 DNA Polymerase Chain Elongation Chain growth - 5’ to 3’ direction 3’ 5’ 5’ 3’

9 DNA Polymerase Chain Elongation

10 Replication Origin Bidirectional Replication from Origin Single origin in Prokaryotes

11 Replication Origins - Replicons Multiple origins in Eukaryotes Replicons

12 Primer Requirement

13 Initiation of DNA Replication - E. coli Initiator protein DnaA binds oriC denaturation DNA helicases unwind DNA DNA primase makes primers

14 Replication Fork - Okazaki Fragments Initial primers Repriming Discontinuous synthesis Lagging strand Continuous synthesis Leading strand 5’ 3’ 5’ 3’ 5’ 3’

15 Activities of DNA Polymerases

16 Types of DNA Polymerases E. Coli DNA polymerase I - single polypeptide DNA 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 primer pol  and pol  - extend primers on leading/lagging strand

17 Joining of Okazaki Fragments

18 DNA Replication in E. coli (Fig. 3.6) Initiation Elongation Primer Removal Ligation

19 DNA Replication in E. coli Figure 3.8. Replisome Model. DNA pol III on lagging-strand template is finishing synthesis of an Okazaki fragment.

20 Telomere Problem Due to: Linear chromosomes & Requirement for RNA primers

21 Resolving the Telomere Problem

22 Rolling Circle Replication

23 Rolling Circle Replication in Phage