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PMID: 372738 Published · ppublish English Journal Article

Plasmid replication functions. III. Origin and direction of replication of a "mini" plasmid derived from R6-5.

Molecular & general genetics : MGG ·Vol. 168 ·No. 1 ·1979-01-05 ·Pages 27-36

Synenki RM, Nordheim A, Timmis KN

Abstract

Replicating DNA molecules of the mini R6-5 plasmid, pKTO71, were purified by equilibrium centrifugation in two successive ethidium bromide-caesium chloride gradients, converted to linear forms by cleavage with either HindIII or BglII restriction endonuclease, and examined in the electron microscope. Determination of the replication fork positions in 65 replicating molecules demonstrated that replication is initiated at a unique location on the plasmid and that it proceeds uni-directionally from this site. The direction of replication is such that the origin-proximal BglII cleavage site is replicated late or, in the case of the parent R6-5 plasmid, is such that the R-determinant region of the molecule is replicated early. The origin of replication, located by these experiments at R6-5 coordinate 98.6 kb, is clearly distinct from that of the R6-5 incompatibility determinant which has been shown to be located on an adjacent PstI-generated DNA fragment whose termini have R6-5 coordinates 96.8 and 97.9 kb. This result indicates that the incompatibility function is not an origin DNA sequence.

MeSH Terms
DNA Replication DNA Restriction Enzymes/metabolism DNA, Bacterial/genetics Escherichia coli/genetics Microscopy, Electron Nucleic Acid Conformation R Factors
Chemicals
DNA, Bacterial DNA Restriction Enzymes
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Synenki R M
Nordheim A
Timmis K N
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36 references, click to expand
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Article Info
Journal
Molecular & general genetics : MGG
Abbr.
Mol Gen Genet
ISSN
0026-8925
DOI
10.1007/BF00267930
Published
1979-01-05
Pages
27-36
Language
English
Region
Germany
NLM ID
0125036
Subset
IM
External Links
PubMed source DOI: 10.1007/BF00267930
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