A Saccharomyces cerevisiae cell‐based quantitative β‐galactosidase assay compatible with robotic handling and high‐throughput screening

Faculty of Life Sciences, University of Manchester, Manchester M13 9PT, UK.
Yeast (Impact Factor: 1.63). 01/2008; 25(1):71-6. DOI: 10.1002/yea.1570
Source: PubMed


Reporter-gene assays that employ the Escherichia coli lacZ gene are ubiquitously employed in biological research. However, we were not able to readily identify a quantitative method that worked reliably with yeast (Saccharomyces cerevisiae) cells and that was compatible with high-throughput screening and robotic liquid handling tools. We have therefore adapted a commercially available assay employing a 6-O-beta-galactopyranosyl-luciferin substrate to provide the required sensitivity with minimal sample handling times. Our assay uses only one-tenth of the reagents suggested by the reagent manufacturer (Promega) for equivalent assays with mammalian cell cultures and produces rapid, sensitive and reproducible analysis with as little as 1 microl yeast cell culture and with < 100 cells. We demonstrate that the assay is compatible with yeast strains generated by the systematic yeast deletion project and functions equally well with genomically integrated or plasmid-encoded lacZ reporters and with cells grown in complex or defined media. The high-sensitivity, miniaturized format reduced sample handling required will make this assay useful for a wide range of applications.

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Available from: Graham D Pavitt, Nov 20, 2014
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