Volume 3

Gene expression / Benjamin Lewin.

  • Lewin, Benjamin.
Date:
1974-1977
Catalogue details

Licence: Attribution-NonCommercial 4.0 International (CC BY-NC 4.0)

Credit: Gene expression / Benjamin Lewin. Source: Wellcome Collection.

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    from 25-60%. When the molecular size of an R factor is known, the number of copies present relative to the host chromosome can be calcu lated by comparing the total radioactive label in the plasmid DNA with that in the bacterial DNA. Isolation in E.coli depends upon the circular state and so any such estimate in this host represents a minimum since it does not take account of any plasmid DNA molecules that may be broken. Estimates of the amount of DNA of the factor R100 and several of its variants made by Nisioka, Mitani and Clowes (1970) show clearly that, in spite of these qualifications, the number of copies of the plasmid is rather low, apparently one or two per host chromosome. This is consis tent with a model in which replication of the plasmid DNA is initiated at a frequency related to that of the bacterial chromosome, limiting the number of copies of plasmid DNA in the cell. This is known as stringent replication control. A somewhat different situation exists in Proteus mirabilis, however, where the number of copies of the plasmid DNA may be very much greater. In the early experiments of Falkow et al. (1966) and Rownd et al. (1966), the satellite bands represented about 8% of the total DNA, which was then thought to correspond to some ten or so copies of the plasmid. Rownd (1969) performed a density transfer analysis of R100 replication in Proteus, analogous to the Meselson-Stahl experiment to demonstrate semiconservative replication (see Volume I of Gene Expression), and could not identify cycles of replication; more than one model for replica tion control is consistent with these data, but the general conclusion that they suggest is that replication is not limited in Proteus to a single synchronous initiation event in each cycle. Another aspect of R factor existence in Proteus is that replication may continue for several hours in the stationary phase, after chromosome replication has ceased; this gen erates a very large number of copies of the plasmid genome, with the result that stationary phase cells contain a much greater proportion of plasmid DNA than cells in exponential growth. (In E.coli, plasmid rep lication halts together with that of the chromosome.) The control of R factor replication in Proteus may be said to be relaxed. The identification of more than one density band of R factor DNA in Proteus raised the possibility that the RTF and r components might be under different types of replication control. Since the r determinant is much smaller tifian the RTF component, the apparent rough equality in the amounts of DNA present in the high and lower density bands suggested that there might be more copies of the r element than of the RTF or composite R factor. Clowes (1972) proposed that in Proteus the r element is under relaxed control whereas the RTF element is under stringent control; the complete R factor is under the control of its RTF component.