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C A Caulcott

Publications and source records attributed to C A Caulcott.

4 recordsLinked to original sources
PubMed1987

A mathematical method for analysing plasmid stability in micro-organisms.

A mathematical model describing the instability of plasmids in micro-organisms has been developed. The model is based on the assumption that the overall causes of plasmid instability are described by the segregational instability of the plasmid, R (i.e. the rate at which plasmid-free cells are generated from plasmid-bearing cells), and the growth rate difference, d mu (i.e. the difference in growth rate between plasmid-free and plasmid-bearing cells). A method for determining the values of R and d mu (accompanied by 95% confidence limits) for any plasmid-bearing micro-organism is described. This method is based on the observation that, depending on the plasmid, various exponential patterns of plasmid instability are observed. The stability of Escherichia coli 1B373(pMG169), where d mu much greater than R, and E. coli RV308(pHSG415), where R much greater than d mu, are analysed in order to demonstrate the method.

Escherichia coli↗
PubMed1987

Investigation of the effect of growth environment on the stability of low-copy-number plasmids in Escherichia coli.

The stability of a low-copy-number plasmid, pHSG415, in Escherichia coli, was investigated in batch and continuous culture. The plasmid was unstable in batch culture, but was significantly stabilized by growth in continuous culture with phosphate, nitrogen or potassium limitation. However, the plasmid was very unstable when grown in continuous culture with sulphate limitation. These results contrast with those obtained with multicopy plasmids such as pBR322, which is particularly unstable in carbon- or phosphate-limited continuous culture. The effect of growth rate on the stability of E. coli(pHSG415) grown in continuous culture with glucose limitation was also investigated. The plasmid was significantly more stable in cells grown at higher growth rates. The segregational instability (R) of the plasmid and the difference in growth rate between plasmid-free and plasmid-bearing cells (dmu) were calculated for each condition using the method of Cooper et al. (accompanying paper: Journal of General Microbiology 133, 1871-1880). It was found that the primary cause of the loss of pHSG415 from the cell population was the segregational instability of the plasmid.

Carbon↗
PubMed1985

Investigation of the instability of plasmids directing the expression of Met-prochymosin in Escherichia coli.

The causes of the instability of a multicopy plasmid, pCT70, which directs the expression of calf prochymosin in Escherichia coli, were investigated. Plasmid pAT153 and its derivative, pCT54, were stable for more than 90 generations in continuous culture with glucose limitation. The multicopy plasmid pCT66, which expressed very low levels of prochymosin due to poor translational efficiency, and low copy number plasmids which efficiently expressed the prochymosin gene, were also stable. These results indicated that high level translation of the recombinant gene was the cause of the instability of pCT70. The maximum specific growth rate of E. coli(pCT70) was reduced by 30% compared with E. coli(pCT66). To fulfil the requirements of a production system, a dual origin plasmid with controllable copy number was developed. Both this plasmid (pMG165) and a derivative which contained the prochymosin gene (pMG168) were stable when maintained at low copy number. When the copy number of plasmid pMG168 was increased by putting replication under the control of the lambda PR promoter and the cI857 temperature sensitive repressor, expression of prochymosin was achieved. This strategy enables large-scale production of prochymosin without the need for antibiotic selection or other methods of preventing plasmid loss.

Chymosin↗