title: Single-step method for β-galactosidase assays in Escherichia coli using a 96-well microplate reader.
abstract: Historically, the lacZ gene is one of the most universally used reporters of gene expression in molecular biology. Its activity can be quantified using an artificial substrate, o-nitrophenyl-ß-d-galactopyranoside (ONPG). However, the traditional method for measuring LacZ activity (first described by J. H. Miller in 1972) can be challenging for a large number of samples, is prone to variability, and involves hazardous compounds for lysis (e.g., chloroform, toluene). Here we describe a single-step assay using a 96-well microplate reader with a proven alternative cell permeabilization method. This modified protocol reduces handling time by 90%.
authors.affiliation: Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK.
authors.forename: Jorrit
authors.initials: J
authors.lastname: Schaefer
authors.affiliation: Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK.
authors.forename: Goran
authors.initials: G
authors.lastname: Jovanovic
authors.affiliation: Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK.
authors.forename: Ioly
authors.initials: I
authors.lastname: Kotta-Loizou
authors.affiliation: Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London SW7 2AZ, UK. Electronic address: m.buck@imperial.ac.uk.
authors.forename: Martin
authors.initials: M
authors.lastname: Buck
mesh_terms: D002478:Cells, Cultured; D057075:Enzyme Assays / Q000295:instrumentation* / Q000379:methods*; D004926:Escherichia coli / Q000166:cytology / Q000201:enzymology*; D001616:beta-Galactosidase / Q000032:analysis / Q000378:metabolism*
chemical_list: D001616:beta-Galactosidase
keywords: B-Galactosidase (Bgal); LacZ; Microplate reader; β-Galactosidase