Effects of Temperature, Agitation, and Donor Strain on Chromosome Transfer in Escherichia coli K-12

Abstract

Recombinant production in Escherichia coli K-12 can be described by three parameters: (i) the distance x of a selected male marker from the donor origin; (ii) the gradient constant k (the probability of interruption of the donor chromosome per unit distance during transfer into a recipient cell); and (iii) the extrapolate number A (the probability that a donor cell will produce a recombinant inheriting the donor marker contiguous with the origin). It is usually assumed that chromosomal distances can be measured by marker entry times, i.e., that the velocity of chromosome transfer v is constant along the chromosome. The dependencies of k, A, x, and v on temperature, agitation during mating, and donor strain were studied. The transfer velocity of the HfrH chromosomal region from the origin to his increases 15-fold between 16 and 43 C, and the chromosomal regions studied have the same temperature dependence that was found for the separate transfer velocities of the O-trp and trp-his regions. These data and radiation studies on chromosome transfer indicate that, at a given temperature, chromosomal transfer velocity varies by less than 10% as the distance of any given region from the origin increases. The gradient constant k is temperature-independent between 20 and 45 C if mating times at different temperatures are inversely proportional to the chromosome velocities; also, k is insensitive to agitation during mating and is not decreased by mating on membrane filters. However, the extrapolate number A is highly temperature-dependent, having its maximum value between 30 and 38 C. These results suggest that the spontaneous interruption of transfer which produces the gradient of transfer is a property of the chromosome itself and not of the fragility of the connection between mating cells.