Recombinant DNA molecules comprising bovine papilloma virus type 1 DNA linked to plasmid DNA are maintained in a plasmidial state both in rodent fibroblasts and in bacterial cells.
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In vitro recombination
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A replicative region of the large conjugative plasmid pHH1457 (incompatibility group HII (IncHII)) was cloned. A 1.4-kbp region, in a stable pSBII14 clone, containing a PolI-independent replicon and determinants for the HII incompatibility phenotype, was selected and characterized. High incompatibility with IncHII plasmids was corroborated. Independent replication of the insert was demonstrated by ligation to an antibiotic resistance cassette. pSBII14 was used as a probe to identify IncHII plasmids from other members of the H complex: IncHI (IncHI1, IncHI2 and IncHI3 subgroups). Hybridization experiments revealed a high homology with the replication region of IncHII plasmids, but not with IncHI1 or IncHI3 plasmid prototypes. Homology with IncHI2 plasmids was observed, suggesting the presence of IncHII-like replicons among this subgroup of plasmids. This is the first report of the characterization of an IncHII plasmid maintenance region.
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The aim of the presented study was to identify the similarity of the plasmid pRK100 RepFIIA replicon (replication region) with similar replicons of other known plasmids of Enterobacteriaceae. For this purpose, within the determined nucleotide sequence of pRK100, theRepFIIA replicon rep genes/regions were identified. The nucleotide sequences of the pRK100 determined rep genes/regions were subsequently compared with the nucleotide sequences of other RepFIIA replicon rep genes/regions deposited in GenBank. Further, the nucleotide divergence between them was calculated. The obtained results clearly demonstrated, that the individual pRK100 rep regions are the same/most similar to rep regions from different plasmids. RepA2 of pRK100 is most similar to repA2 of pCP301, pINV_F6_M1382, pWR501 and R1, copA is the same as copA of plasmids pC15-1a and R100, repA6 of pRK100 is the same as repA6 in plasmids pC15-1a, pCP301, pINV_F6_M1382, pWR501, R1 and R100, repA1 is most similar to repA1 of the plasmid p1658/79, and repA4 of pRK100 is most similar to repA4 of pC15-1a. Hence, the composition of the pRK100 RepFIIA replicon is mosaic and unique among the plasmids.
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Objective To observe the eliminating effect of dual-replicon plasmids on resistant plasmids.Methods The dual-replicon plasmids pKT230-oriV were constructed and transferred into bacteria with resistant plasmids pRK290through the transformation and conjugational transfer pathways.The eliminating effect of dual-replicon plasmids on resistant plasmids was observed.Results The bacteria were cultured for 5generations after the transfer of dual-replicon plasmids,and resistant plasmids pRK290in bacteria were eliminated.Conclusion The conjugational transfer of dual-replicon plasmids is a pathway to eliminate resistant plasmids.
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Two replicons were isolated independently from different IncHI1 plasmids. One was isolated from R27, and a second was isolated from pIP522. We demonstrate, by DNA-DNA hybridization experiments, that these maintenance regions are different and that they are specific to, and carried by, all IncHI1 plasmids tested. In view of this specificity we decided to designate the replicon isolated from R27 as RepHI1A and the replicon isolated from pIP522 as RepHI1B. These two autoreplicative regions are not related to a third replicon present in all IncHI1 plasmids that bears homology with RepFIA and that expresses the characteristic incompatibility of IncHI1 subgroup plasmids toward F factor (D. Saul, D. Lane, and P. L. Bergquist, Mol. Microbiol. 2:219-225, 1988; D. E. Taylor, R. W. Hedges, and P. L. Bergquist, J. Gen. Microbiol. 131:1523-1530, 1985). These results demonstrate that all IncHI1 plasmids tested contain at least three replicons. An incompatibility (Inc) region that hybridizes specifically to all the IncHI1 plasmids was previously isolated (M. Couturier, F. Bex, P. L. Bergquist, and W. K. Maas, Microbiol. Rev. 52:375-395, 1988). Although this Inc locus is not located in an autoreplicative region of IncHI1 plasmids, we observed that this locus stabilizes a low-copy-number replicon. This Inc locus is probably a component of an active partition locus involved in the maintenance of IncHI1 plasmids. The nucleotide sequence of the Inc region contains direct repeats of 31 bp. In addition, this incompatibility determinant hybridizes specifically with IncHI1 plasmids but expresses incompatibility toward plasmids of both IncHI subgroups (IncHI1 and IncHI2). In this communication, we present the mapping of these maintenance elements on the R27 genome.
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