DNA replication initiates at multiple sites on plasmid DNA in Xenopus egg extracts

H. M. Mahbubani, T. Paull, J. K. Elder, J. J. Blow

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    112 Citations (Scopus)


    Cell-free extracts of Xenopus eggs will replicate plasmid DNA molecules under normal cell cycle control. We have used the neutral/neutral 2-D gel technique to map the sites at which DNA replication initiates in this system. Three different plasmids were studied: one containing the Xenopus rDNA repeat, one containing single copy Xenopus genomic DNA, and another containing the yeast 2 microns replication origin. 2-D gel profiles show that many potential sites of initiation are present on each plasmid, and are randomly situated at the level of resolution of this technique (500-1000 bp). Despite the abundance of sites capable of supporting the initiation of replication, pulse-chase experiments suggest that only a single randomly situated initiation event occurs on each DNA molecule. Once initiation has taken place, conventional replication forks appear to move away from this site at a rate of about 10nt/second, similar to the rate observed in vivo.
    Original languageEnglish
    Pages (from-to)1457-1462
    Number of pages6
    JournalNucleic Acids Research
    Issue number7
    Publication statusPublished - 11 Apr 1992


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