Electrochemical evaluation of cellular physiological status under stress in Escherichia coli with the rpoS-lacZ reporter gene.
We developed an electrochemical detection method for evaluating cellular physiological status based on the stringent response as a means to monitor cell viability. A reporter plasmid was constructed by inserting the beta-galactosidase gene (lacZ) under the control of the rpoS promoter, and then used to transform E. coli cells. Electrochemical responses from the products catalyzed by beta-galactosidase expressed by these E. coli cells were detected using the chronoamperometric technique in a nondestructive manner. Comparisons of response currents between the relA-positive strain and relA-negative strain revealed that increases in these currents were caused by the stringent response due to the stressful alcoholic environment, and thus as a model of stressful cultivating conditions. The current was proportional to the beta-galactosidase activity assayed by a conventional method that required the destruction of cells. The cellular physiological status, which depends on the stringent response as a viability marker, therefore, could then be evaluated online with a current using the rpoS-lacZ reporter gene in the relA-positive strain without pretreatment.
Funabashi
Hisakage
H
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Tokyo, Japan.
Ishikawa
Miyuki
M
Mie
Masayasu
M
Takahashi
Fumio
F
Yanagida
Yasuko
Y
Aizawa
Masuo
M
Kobatake
Eiry
E
D001426:Bacterial Proteins / Q000235:genetics*; D015374:Biosensing Techniques; D004563:Electrochemistry; D004784:Environmental Monitoring / Q000379:methods*; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D000431:Ethanol / Q000494:pharmacology; D017930:Genes, Reporter; D007763:Lac Operon / Q000235:genetics; D012808:Sigma Factor / Q000235:genetics*; D001616:beta-Galactosidase / Q000032:analysis* / Q000235:genetics
D001426:Bacterial Proteins; D012808:Sigma Factor; C069567:sigma factor KatF protein, Bacteria; D000431:Ethanol; D001616:beta-Galactosidase