1. Avery, McCarty, and MacLeod Experiment What were their conclusion? CarbohydratesLipidsProteinsRNADNA Mice were given deadly bacteria with enzymes that destroyed …

2. S-Type Carbohydrates Destroyed S-Type Lipids Destroyed S-Type Proteins Destroyed S-Type RNA Destroyed S-Type DNA Destroyed Conclusion: DNA was the transforming factor! DNA is the molecule of inheritance

3. Learning Outcomes Content Explain which experiment demonstrated that DNA was the molecule of inheritance Investigate the theories of replication Process Conclusion to experimentation Benefit Explain how experimentation can make leaps forward in our understanding.

4. 3 proposed theories of how DNA replicates.... Any ideas to test these? Take an educated guess of which model and WHY!

5. N 15 sole source of nitrogen for many generations Collect and analyze DNA Cell Transfer Transfer cells to N 14 and Allow cells to divide only once Collect and analyze DNA Allow cells to divide a second time Collect and analyze DNA Prediction Prediction Prediction Conservative All heave labeled DNA Semi conservative Not Relevant Conservative First generation ½ labeled with only heavy label ½ labeled with N 14 Semi conservative All DNA hybrids N 15 /N 14 Conservative Only one heavy DNA 3 DNA N 14 Semi conservative ½ DNA N 15 ½ DNA N 14

6. Meselson – Stahl Experiment To answer this question Meselson and Stahl exploited the differences between N 14 and N 15 –N 15 is a heavy isotope with one extra neutron –The difference in mass creates a difference in density –This difference in density can be seen when centrifugation techniques are employed

8. DNA (helicase) unwinds to form 2 templates (RNA) primer is then bound to the starting DNA site. Begin of replication, an enzyme called DNA polymerase binds to the (RNA) primer, which indicates the starting point for the replication. Occurs at several locations on DNA strandDNA polymerase DNA polymerase can only synthesize new DNA The DNA polymerase can only travel on one side of the original strand without any interruption (3’ to 5’). Since the DNA replication on the lagging strand is not continuous, the replicated DNA is fragmented with fragments added by the enzyme ligase. Semi-conservative replication!

9. Activity – Artists Easel Now draw the parts for each part I have described.

10. Self Check Compare what I’ve drawn to your own diagram/pictures....

11. DNA Replication From this discovery the mechanism of how DNA replicates was proposed... 11 Begins as DNA unwinds (helicase)Begins as DNA unwinds (helicase) Two strands open forming Replication Forks (Y-shaped region)Two strands open forming Replication Forks (Y-shaped region) New strands grow at the forksNew strands grow at the forks ReplicationFork Parental DNA Molecule 3’ 5’ 3’ 5’

12. 12 DNA Replication DNA polymerase can only add nucleotides to the 3’ end of the DNADNA polymerase can only add nucleotides to the 3’ end of the DNA This causes the NEW strand to be built in a 5’ to 3’ directionThis causes the NEW strand to be built in a 5’ to 3’ direction RNAPrimer DNA Polymerase Nucleotide 5’ 3’ Direction of Replication

13. 13 DNA Replication As the 2 DNA strands open at the origin, Replication Bubbles formAs the 2 DNA strands open at the origin, Replication Bubbles form Meaning DNA replication occurs at several locationsMeaning DNA replication occurs at several locations Bubbles copyright cmassengale

14. 14 Synthesis of the New DNA Strands The Leading Strand single strandThe Leading Strand is synthesized as a single strand from the point of origin toward the opening replication fork RNAPrimer DNA Polymerase Nucleotides 3’5’

15. 15 Synthesis of the New DNA Strands The Lagging Strand is discontinuouslyThe Lagging Strand is synthesized discontinuously against overall direction of replication This strand is made in MANY short segments It is replicated from the replication fork toward the origin RNA Primer Leading Strand DNA Polymerase 5’ 3’ Lagging Strand 5’ 3’

16. 16 Lagging Strand Segments (Okazaki Fragments) - lagging strand(Okazaki Fragments) - series of short segments on the lagging strand Must be joined together by an enzyme LigaseMust be joined together by an enzyme Ligase Lagging Strand RNAPrimerDNAPolymerase 3’ 5’ Okazaki Fragment copyright cmassengale

17. 17 Joining of Okazaki Fragments The enzyme Ligase joins the Okazaki fragments together to make one strandThe enzyme Ligase joins the Okazaki fragments together to make one strand Lagging Strand Okazaki Fragment 2 DNA ligase DNA ligase Okazaki Fragment 1 5’ 3’

18. 18 Replication of Strands Replication Fork Point of Origin

19. 19 Question: What would be the complementary DNA strand for the following DNA sequence? DNA 5’-CGTATG-3’ copyright cmassengale

20. 20 Answer: DNA 5’-CGTATG-3’ DNA 3’-GCATAC-5’ copyright cmassengale

21. Activity Add a series of diagrams/cartoons which explain the terms

22. 22 Proofreading New DNA DNA polymerase initially makes about 1 in 10,000 base pairing errorsDNA polymerase initially makes about 1 in 10,000 base pairing errors Enzymes proofread and correct these mistakesEnzymes proofread and correct these mistakes The new error rate for DNA that has been proofread is 1 in 1 billion base pairing errorsThe new error rate for DNA that has been proofread is 1 in 1 billion base pairing errors

23. 23 Semiconservative Model of Replication Idea presented by Watson & CrickIdea presented by Watson & Crick TheThe two strands of the parental molecule separate, and each acts as a template for a new complementary strand New DNA consists of 1 PARENTAL (original) and 1 NEW strand of DNA Parental DNA DNA Template New DNA copyright cmassengale