The parB region of plasmid R1 encodes two genes, hok and sok, which are required for the plasmid‐stabilizing activity exerted by parB. The hok gene encodes a potent cell‐killing factor, and it is regulated by the sok gene product such that cells losing a parB‐carrying plasmid during cell division are rapidly killed. Coinciding with death of the host cell, a characteristic change in morphology is observed. Here we show that the killing factor encoded by the hok gene is a membrane‐associated polypeptide of 52 amino acids. A gene located in the Escherichia coli relB operon, designated relF, is shown to be homologous to the hok gene. The relF gene codes for a polypeptide of 51 amino acids, which is 40% homologous to the hok gene product. Induced overexpression of the hok and relF gene products results in the same phenomena: loss of cell membrane potential, arrest of respiration, death of the host cell and change in cell morphology. The parB region and the relB genes were cloned into unstably inherited oriC minichromosomes. Whereas the parB region also conferred a high degree of genetic stability to an oriC minichromosome, the relB operon (with relF) did not; therefore the latter does not appear to ‘stabilize’ its replicon (the chromosome). The function of the relF gene is not known.

The potential risks of unintentional releases of genetically modified organisms, and the lack of predictable behavior of these in the environment, are the subject of considerable concern. This concern is accentuated in connection with the next phase of gene technology comprising deliberate releases. The possibilities of reducing such potential risks and increasing the predictability of the organisms are discussed for genetically engineered bacteria. Different approaches towards designing disabled strains without seriously reducing their beneficial effects are presented. Principally two types of strain design are discussed: actively contained bacteria based on the introduction of controlled suicide systems, and passively contained strains based on genetic interference with their survival under environmental-stress conditions.

The two replication origins of plasmid pUB110 have been characterized. The site of initiation of DNA replication at the plus origin was mapped to within an 8-base-pair sequence. DNA synthesis initiated at the origin was made to terminate precociously in an inserted sequence of 18 base pairs that is homologous to a sequence in the origin. This suggests that pUB110 replicates as a rolling circle. The minus origin of plasmid pUB10 has been characterized, and the minimal sequence required for function has been determined. As with other minus origins, activity is orientation specific with respect to the direction of replication. Its activity is sensitive to rifampin in vivo, suggesting that RNA polymnerase catalyzes single-strand to double-strand conversion. Unlike all other plasmids of gram-positive bacteria thus far described, the pUB110 minus origin is functional in more than one host.

Summary When bacterial cells are subjected to a strong selective pressure it often induces specific mutations. Here a model is considered in which errors are introduced at random in one of the strands of the DNA molecule: a nick in one of the strands can initiate strand displacement rendering a region of the chromosome single‐stranded. Upon conversion back to double‐stranded DNA there is a certain probability of introducing errors creating a heteroduplex. If an error results in the production of an mRNA molecule encoding a product which provides a selective advantage, growth will be stimulated and the mutation can be immortalized by chromosomal replication. Otherwise, the error can be corrected by the DNA ‘proofreading’ enzymes.

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Mutations ofEscherichia coli from sensitivity to nalidixic acid resistance were studied by fluctuation analysis.The mutant distributions in replicate cultures were not significantly affected either by the age of the carbon-starved preculture used for inocula or by the inoculum size. The data from 23 fluctuation tests (48 cultures each) were pooled. The mean number of mutations per culture was estimated to be 0.71 from the fraction of cultures without mutants or 0.74 and 0.77 by maximum-likelihood estimation based on the two models under consideration. When the pooled data were compared with the theoretical expectations, the fits were unsatisfactory (P < 0.005). The lack of fit was caused mainly by too high a frequency of cultures with between 17 and 32 mutants and too high a frequency of cultures with more than 128 mutants. Possible reasons for the lack of fit and its implications with respect to estimation of mutation rates from fluctuation tests are discussed.The statistical analysis of bacterial mutations came into prominence with the publication of Luria and Delbruck (16); the system they studied was that of resistance to bacteriophage Ti in Escherichia coli. When a culture of sensitive E. coli is plated in the presence of an excess of the phage, although virtually all the bacteria are killed, a small number survive and give rise to colonies on the plate. The problem analyzed by Luria and Delbruck was whether the survivors were the result of mutations that occurred prior to the plating, i.e., during the growth of the culture, or whether they occurred after plating, i.e., as a direct consequence of the bacteria being exposed to the phage. These two principal conceptions may be labelled the pre-and postadaptive theories, respectively (7). Luria and Delbruck studied the problem using mathematical models. If the mutation occurred postadaptively, the distribution of mutants in a number of replicate (parallel) cultures should be a Poisson distribution, provided that all bacteria have a small, finite probability of mutating to become resistant after the exposure to the phage. The preadaptive mutation theory leads to a quite different prediction. Although the replicate cultures are the same with respect to the total size of population, some may have experienced a mutation at an early generation and thereby yield a large number of mutants, while with others, a mutation may have occurred just prior to the plating so that only one or a few mutants are present in these cultures. The difference in the predictions of the pre-and postadaptive mutation theories could therefore be reduced to a prediction of the variance of the number of mutants in replicate cultures: the postadaptive theory predicted that the variance would equal the mean, whereas the preadaptive mutation theory predicted that the variance would be much larger, owing to a few cultures yielding a large number of mutants.Luria (45)45932809. from each culture. They did not correct for sampling error, nor did they deduce the theoretical distribution expected fr...