Insertion of an R1 plasmid into the origin of replication of the E.Two-dimensional gel electrophoresis analysis showed that Mcm10p is required not only for the initiation of DNA synthesis at replication origins but also for the smooth passage of replication forks at origins.) copies of the replication origin, as determined by flow cytometry. ) nucleoids, as determined by 4',6-diamidino-2-phenylindole staining, and 2n (n = 1, 2, 3. We show here that the overwhelming majority of nucleate cells contain 2n (n = 0, 1, 2.After treatment with alkaline phosphatase, T antigen had a severalfold higher affinity for the specific binding sites in the viral genomic control region, in particular, for binding site II in the origin of replication.Utilizing nitrocellulose filter binding, DNase I footprinting, and gel mobility shift assays, we observed that ATP increased Py TAg binding to DNA fragments containing either all Py TAg-binding sites (whole origin) or those sites within (core origin) or adjacent to (early) the origin of replication.The intracellular locations of oriC and oriR1, the replication origins of the chromosome and plasmid R1, respectively, were visualized by fluorescence in situ hybridization (FISH) in exponentially growing populations of Escherichia coli.Īnalytical, diagnostic and therapeutic context of Replication Origin The rate of excision repair of UV-induced pyrimidine dimers (PDs) was measured at specific sites in each strand of a yeast minichromosome containing an active gene ( URA3), a replication origin (ARS1), and positioned nucleosomes.Based on binding of plasmid DNA fragments, dnaA protein recognizes in addition to oriC a number of specific sites: within or near the replication origins of pSC101, pBR322, and ColE1 within the regulatory regions of the dnaA and "X-protein" genes and in IRL-Tn5.The rest of the viral genome including the origin of replication and T antigen binding region is present and colinear with SV40 DNA, except for an internal repeat of about 1750 bp located between nucleotides 27.In the postreplicative state, genomic footprints closely resemble those produced in vitro by the purified ORC and ABF1 proteins, indicating that the binding of these proteins to replication origins is not sufficient to drive the initiation of DNA replication.coli chromosome are not stably inherited through cell division, but can be stabilized by joining with a particular segment of F plasmid that presumably provides the partition function. Plasmids that replicate using the replication origin ( oriC) of the E.We determined the nucleotide sequence of RNA synthesized in vitro by Escherichia coli RNA polymerase at the complementary-strand replication origin on the single-stranded viral DNA of bacteriophages f1 and IKe (ori-RNA) by using chain-terminating ribonucleoside triphosphate analogs.After ligation to a tetracycline resistance cassette, fragments containing a bacterial origin of replication from the insertion are recovered in Escherichia coli because they replicate autonomously.Both yeast and human RP-A bind preferentially to the pyrimidine-rich strand of a homologous origin of replication: the ARS307 or the simian virus 40 origin of replication, respectively.The replication origin of Escherichia coli has been cloned on a nonreplicating DNA fragment coding for ampicillin resistance.To study recombination between short homologous sequences in Escherichia coli we constructed plasmids composed of the pBR322 replicon, M13 replication origin and a recombination unit inserted within and inactivating a gene encoding chloramphenicol resistance.Ĭhemical compound and disease context of Replication Origin Mechanism of phage Mu-1 integration: nalidixic acid treatment causes clustering of Mu-1-induced mutations near replication origin.After transfection with chrysotile, a minor percentage of introduced plasmid DNA bearing a simian virus 40 origin of replication replicates after 24 hr.Both the linear cloned viral DNA and a hybrid composed of mutant Bam HI (0.58) to Bgl I (0.72) 750 bp fragment (containing the origin of replication) ligated to the complementary wild-type 4.5 kb fragment are able to multiply on PCC4 embryonal carcinoma cells.These results demonstrate that adenovirus encodes a novel DNA polymerase that is required for priming of DNA synthesis at the origin of replication.A 16 bp BgI II fragment was deleted in vitro from the minimal origin of replication of the Escherichia coli chromosome, oriC, and was replaced by a 10 kb R1 miniplasmid, pKN1562, containing the basic R1 replicon and a kanamycin resistance gene.
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