lecture 06 – dna replication based on chapter 3 dna replication copyright © 2010 pearson...
TRANSCRIPT
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Lecture 06 – DNA ReplicationBased on Chapter 3 DNA Replication
Copyright © 2010 Pearson Education Inc.
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1. Semiconservative DNA Replication
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2a. The Meselson–Stahl Experiment
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2b. The Meselson–Stahl Experiment
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2c. The Meselson–Stahl Experiment
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3. DNA Polymerase (Kornberg Enzyme)4. DNA Polymerase I
• First isolation of an enzyme involved in DNA replication was in 1955. Arthur Kornberg won the 1959 Nobel Prize in Physiology or Medicine for this work.
• Kornberg isolated DNA polymerase I.• Availability of DNA Pol I allowed characterization of DNA
synthesis in vitro. Five components are needed:– Radioactively labeled dNTPs (all four: dATP, dGTP, dTTP,
and dCTP).– DNA polymerase I.– E. coli DNA (template).– Primer DNA.– Magnesium ions (Mg++).
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5. Roles of DNA Polymerases
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6. Molecular Model of DNA Replication
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7. Initiation of Replication
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8a. Semidiscontinuous DNA Replication
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8b. Semidiscontinuous DNA Replication
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8c. Semidiscontinuous DNA Replication
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8c. Semidiscontinuous DNA Replication
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8d. Semidiscontinuous DNA Replication
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8e. Semidiscontinuous DNA Replication
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9. DNA Replication in Eukaryotes
10. Replicons
DNA replication is very similar in both prokaryotes and eukaryotes, except that eukaryotes have more than one chromosome.
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11. Initiation of Replication
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12. Eukaryotic Replication Enzymes
• Enzymes of eukaryotic DNA replication aren’t as well characterized as their prokaryotic counterparts.
• Fifteen DNA polymerases are known in mammalian cells:• Three DNA polymerases are used to replicate nuclear DNA.
– Pol a (alpha) extends the 10-nt RNA primer by about 30 nt.– Pol d (delta) and Pol ´ (epsilon) extend the RNA/DNA
primers, one on the leading strand and the other on the lagging (it is not clear which synthesizes which).
• Other DNA Pols replicate mitochondrial or chloroplast DNA, or they are used in DNA repair.
• Primer removal differs from that in prokaryotes. – Pol d continues extension of the newer Okazaki fragment,
displacing the RNA and producing a flap that is removed by nucleases, thus allowing the Okazaki fragments to be joined by DNA ligase.
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13a. Replicating the Ends of Chromosomes
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13b. Replicating the Ends of Chromosomes
• Most eukaryotic chromosomes have short, tandemly repeated sequences at their telomeres
• Telomerasecontains both protein and RNA • 11-bp RNA sequence used to synthesize
the new telomere repeat DNA. • Telomerase functions as reverse
transcriptase.• After telomerase adds telomere sequences,
chromosomal replication proceeds in the usual way.
• Telomere length may vary• Loss of telomerase activity results in limited
rounds of cell division before the cell death.
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14. Assembling Newly Replicated DNA into Nucleosomes