A rapid reporter system using GFP as a reporter protein for identification and screening of synthetic stationary-phase promoters in Escherichia coli.
To develop a rapid reporter system for the screening of stationary-phase promoters in Escherichia coli, the expression pattern of the green fluorescent protein (GFP) during bacterial cultivation was compared with that of the commonly used beta-galactosidase. Using GFP with enhanced fluorescence, the expression pattern of both reporter systems GFP and beta-galactosidase were similar and showed a typical induction of gene activity of the reporter genes, i.e. increase of expression at the transition from exponential to stationary phase. The expression was affected by the culture medium, i.e. in contrast to the complex medium (LB medium), the stationary-phase specific induction was only observed in synthetic medium (M9) when amino acids were added, whereas there was generally no induction in MOPS medium. To develop a rapid screening method on agar plates for stationary-phase promoters, a photographic approach was used, continued with computational image treatment. A screening method is presented which enables an on-line monitoring of gene activity.
Miksch
G
G
Lehrstuhl für Fermentationstechnik, Technische Fakultät, Universität Bielefeld, Germany. gmi@fermtech.techfak.uni-bielefeld.de
Bettenworth
F
F
Friehs
K
K
Flaschel
E
E
Saalbach
A
A
Nattkemper
T W
TW
D001709:Biotechnology / Q000379:methods; D019295:Computational Biology; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D007091:Image Processing, Computer-Assisted; D011401:Promoter Regions, Genetic; D013997:Time Factors; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D003470:Culture Media; D029968:Escherichia coli Proteins; D049452:Green Fluorescent Proteins; D001616:beta-Galactosidase