Fluorescence-based detection of lacZ reporter gene expression in intact and viable bacteria including Mycobacterium species.
A variety of fluorescein di-beta-D-galactopyranoside (FDG)-based substrates were evaluated for measuring beta-galactosidase expression in bacteria. One substrate, 5-acetylamino-FDG (C2FDG), performed well in all bacteria tested, including the slow growing mycobacterium, Mycobacterium bovis BCG. The sensitivity of C2FDG in intact, viable BCG was similar to that of o-nitrophenyl-beta-D-galactopyranoside in cell lysates when used to measure lacZ reporter gene activity. C2FDG was approximately 70-fold more sensitive than green fluorescent protein (GFP) in BCG when assayed in a fluorescence plate reader, and comparable to GFP when measured by flow cytometry. These assays provide an important new alternative for the rapid measurement of reporter gene expression in viable bacteria.
Rowland
B
B
David Axelrod Institute, Wadsworth Center, New York State Department of Health, P.O. Box 22002, 120 New Scotland Ave, Albany, NY 12208, USA.
Purkayastha
A
A
Monserrat
C
C
Casart
Y
Y
Takiff
H
H
McDonough
K A
KA
D001419:Bacteria / Q000235:genetics*; D005452:Fluoresceins; D005453:Fluorescence; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D007763:Lac Operon; D008164:Luminescent Proteins / Q000235:genetics; D009161:Mycobacterium / Q000235:genetics*
D005452:Fluoresceins; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins