Pattern of Replication of Various Genes in Exponential and Synchronized Cultures of Escherichia coli

Abstract

Early experiments to locate the origin of replication on the E. coli chromosome used a variety of techniques: pulse mutagenesis with nitrosoguanidine, enzyme induction or transduction with bacteriophage PI. They provided the general conclusion that there was a fixed origin of replication, in the lower left quadrant of the E. coli genetic map, and that replication proceeded uni-directionally in a clockwise manner. Some of the data obtained with PI transduction could be viewed however as indicating bi-directional replication (Caro and Berg, 1968). Recently, we have described a more precise method for determining origin and direction of chromosome replication in E. coli (Bird et al., 1972). The origin and the direction of replication were defined in two ways: the gradient of marker frequency in exponential cultures and the sequence of marker replication after partial synchronization. Gene frequency and sequence of replication were assayed by DNA-DNA hybridization using only two markers. The first, the prophage lambda, is fixed and the second, the prophage Mu-1, is moveable. We have constructed an isogenic series of strains, each lysogenic for prophage lambda and for Mu-1 integrated into a different chromosomal site. Mu-1 often integrates within a gene, thus making the identification of its location quite simple.