Monitoring of genes that respond to process-related stress in large-scale bioprocesses

in large-scale aerobic fed-batch processes, cells are exposed to local zone s of high glucose concentrations that can also cause local oxygen limitatio ns at high cell densities. The mRNA levels of four stress genes (clpB, dnaK , uspA, and proU) and three genes responding to oxygen limitation or glucos e excess (pfl, frd, and ackA) were investigated in an industrial 20-m(3) Es cherichia coli process and in a scale-down reactor with defined high-glucos e and low-oxygen zones. The mRNA levels of ackA and proU were high during t he batch growth phase, but declined drastically when glucose became limited , whereas the mRNA levels of the other stress genes were relatively constan t throughout the process. In the industrial-scale reactor, the stress gene mRNA levels were, in most cases, highest in the middle part and at the top of the reactor, where the substrate was fed. Cells passing through the high glucose zone of the scale-down reactor had elevated mRNA levels for the ox ygen limitation genes and had also elevated heat-shock gene mRNA levels. Bo th responses to stress occurred within seconds. The approach presented in t his study offers a tool for monitoring process-related changes in the trans criptional regulation of genes. (C) 1999 John Wiley & Sons, Inc.