Lecture6 Note

Lecture6 Note - Mon. 8.17.09 Lecure 6: DNA Replication I....

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Mon. 8.17.09 Lecure 6: DNA Replication I. The nature of DNA Replication DNA replication occurs during S-phase of the cell division. o Mitosis: when the chromosomes condense. (that’s what we usually see in a cell) However, this takes only 10% (time-wise) of the whole cell division. o S-phase: DNA synthesis; chromosome duplication. Takes up to 90% of time in the cell division process. o In between S-phase and the mitosis, there are G1 and G2, which serves as the checkpoint. What is the nature of DNA Replication? o Hypothesis1: conservative Parental strand is conservative. A brand new DNA strand is created. o Hypothesis2: semiconservative One of the parental strands is passed onto the new strand. 1 st generation consist of one new strand and one parental strand. Parental strand is used as ‘template’ o Hypothesis3: dispersive Most unlikely situation. 1 st generation consist of segments from parental and new strand. o Isotopes: same atom with different # of neutrons. o To analyze the change of density of DNA isolated, use density gradient centrifugation to separate DNA. o The DNA strands are able to have different density since parental DNA would be exposed to N15. Once the1st generation is born, it’d be exposed to N14. This way, we would be able to use density gradient centrifugation. Origin of Replication: Where will replication start in a genome? o Replication origin (or Ori, replicon) is a specific sequence in the genome, in which the dsDNA is first opened up for replication. o Bacteria has circular chromosome. o Prokaryote – most prokayotes have one replicon per circular genome. Plasmid and viruses also have only one replicon per genome. But prokaryotic sequencing projects have revealed that some prokaryotes have linear chromosomes and multiple replicons. A eukaryotic genome has many origins of replication per chromosome.
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o The replication can go in both directions. o The genome of eukaryotes is much larger than prokaryotes. Takes too long to completely replicate entire genome in a cell with one origin of replication. o Issue is resolved if have multiple origins of replication Features of Replication Origin o They are short repeats of DNA sequences that are rich in A/T. In order to replicate, DNA structure must be denatured. Since A/T is easier to break than G/C because of # of hydrogen bonds in between the nucleotides, DNA sequence with rich A/T content is used. E.coli OriC is about 248bp; one core sequence is TGTGGATAA Yeast have ARSI (autonomous replication sequence 1); core sequence: A/ TTTTAA/GTTTA/T o They are bound by specific ori-binding proteins (e.g DNA in A in E. coli or ORC in SSV40). The Origin-binding proteins will bind to additional enzymes in order to start replication. Unidirectional vs. Bidirectional Replication
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Lecture6 Note - Mon. 8.17.09 Lecure 6: DNA Replication I....

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