Cell cycle characteristics and changes in membrane potential during growth of Escherichia coli as determined by a cyanine fluorescent dye and flow cytometry

A preliminary study on the optimization of the DiOC 6(3) staining technique using flow cytometry for evaluating the membrane potential of Escherichia coli showed that DiOC 6(3) was an adequate membrane potential probe. Flow cytometry was used to determine the growth curve by cell direct counts, to examine cell cycle characteristics by measuring DNA content using Hoechst 33342, and to observe changes in membrane potential using DiOC 6(3) of E. coli in optimum culture conditions. The membrane potential increased abruptly from lag phase and the maximum value occurred before the mid-exponential phase at the end of 1 h 30 min of culture. At the same time, replication activity, as determined by measuring the number of DNA replication origins, reached its maximum. After the mid-exponential phase, while generation time was 23 min, membrane potential and DNA replication decreased steadily until the stationary phase, with cells using the replicated chromosome stocks. The rapid increase of membrane potential as early as the lag phase until just before the mid-exponential phase suggested that metabolic activity was maximum during this period to respond to energy supplies that were necessary, in part, for DNA replication.