GDCB 511-4-13-09

GDCB 511-4-13-09 - GDCB 511 13 April 2009 DNA Replication...

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GDCB 511: 13 April 2009, DNA Replication in Prokaryotes The basic unit of replication is the replicon in which all the DNA replicated from a single origin. Origin provides site of initiation of replication. In DNA replication, replicating forks represent sites of DNA replication
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Mechanisms of chromosomal DNA replication in bacteria and bacteriophage Basic Features: 1. Template copied via complementary basepairing (proposed by Watson &Crick, 1953) 2. Semi-Conservative: Meselson and Stahl, 1958. Density-gradient ultracentrifugation of density- labeled DNA. Figure 20.1
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The Meselson Stahl Experiment Figure 20.3
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DNA Replication Models
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Lagging strand synthesis is discontinuous Okazaki fragments: labeled nucleotides are first incorporated into very short fragments. Wild type DNA ligase mutant Figure 20.6
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3. 5’ to 3’ synthesis: This is the direction of synthesis by all known DNA polymerases. dNTP’s are added to the 3’OH of the growing chain. This polarity of replication of the anti-parallel DNA strands leads to the requirement of leading and lagging strands of DNA synthesis: Basic Features of DNA Replication (continued): Leading strand: Continuous synthesis, in 5’ to 3’ direction. Lagging strand: Discontinuous synthesis in discrete fragments (Okazaki fragments). Each fragment is initiated by a short (10 – 12 bp) RNA primer. DNA polymerase then adds DNA to the 3' OH of the RNA primer. RNA primer removed, gaps filled, and nicks ligated.
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Semidiscontinuous Replication DNA replication in E. coli is semidiscontinuous One strand is replicated continuously in the direction of the movement of the replicating fork The other strand is replicated discontinuously as 1-2 kb Okazaki fragments in the opposite direction This allows both strands to be replicated in the 5’ 3’-direction
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Initiated by a primer: DNA polymerase cannot initiate synthesis de novo; it adds nucleotides only to an existing 3' hydroxyl group on an RNA or DNA primer.
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GDCB 511-4-13-09 - GDCB 511 13 April 2009 DNA Replication...

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