mcb450-23s07 - Office Hours Tuesday, April 17, 1:00-2:30...

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Office Hours Tuesday, April 17, 1:00-2:30
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Lecture 23 DNA Replication and Repair
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Models for DNA Replication Conservative Semiconservative Dispersive
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Grew E. coli for many generations in a medium containing a heavy, stable, isotope of nitrogen, 15 N. The bases of DNA would then contain some 15 N and DNA’s so labeled would have greater density then 14 N containing DNA. Transferred cells to an 14 N containing medium. Followed DNA replication for a number of generations in the light medium. Carried out density-gradient ultracentrifugation and found a pattern consistent with semi-conservative replication.
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Meselson & Stahl Experiment
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Semiconservative Replication
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Bidirectional Replication Add radioisotope to cells replicating DNA, then carry out autoradiography.
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Properties of DNA Polymerases Read template strand in 3′-5′ direction. Synthesize new DNA in 5′-3′ direction. This poses a problem as DNA strands open. Solution is that one strand is synthesized continuously and the other discontinuously.
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Semidiscontinuous Replication
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E. coli DNA Polymerases
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E. coli DNA Polymerase I First DNA polymerase discovered Catalyzes the following reaction: n(dATP,dTTP,dGTP,dCTP) = DNA + nPPi Requirements: Template: Complementary strand Primer: Can only extend existing chains Attaches incoming nucleotide to the free 3′-OH group of the primer Reads template strand in 3′→5′ direction Not the major enzyme involved in DNA replication, but necessary for replication.
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E. coli DNA Polymerase I: Other Activities Serves as 3′→5′ exonuclease (3′ exonuclease) Important in proofreading. A newly inserted nucleotide can be exised if it is mis-paired. Think about the backspace key on a computer. Serves as 5′→3′ exonuclease (5′ exonuclease)
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This note was uploaded on 04/07/2008 for the course MCB 450 taught by Professor Mintel during the Fall '07 term at University of Illinois at Urbana–Champaign.

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mcb450-23s07 - Office Hours Tuesday, April 17, 1:00-2:30...

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