"One plate/three-reporter" assay format for the detection and validation of yeast two-hybrid interactions.
Biotechniques
; 37(5): 840-3, 2004 Nov.
Article
em En
| MEDLINE
| ID: mdl-15560140
ABSTRACT
We describe a novel assay format for the Gal4-based yeast two-hybrid-system, in which the readout from three different reporter genes is measured sequentially in a single microplate. Activation of the URA3, MEL1, and lacZ reporters in response to a protein-protein interaction is monitored by measuring sequentially (i) growth in medium lacking uracil, (ii) alpha-galactosidase activity, and (iii) beta-galactosidase. The data thus generated permit elimination of many false positive signals and provide a preliminary measurement of reporter activation-strength that may be confirmed by further analysis. The assay procedure is inexpensive and requires few liquid-handling steps. It is appropriate for automated high-throughput interaction mating assays, validation of putative interactor strains and hybrid-protein self-activator tests.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Leveduras
/
Proteínas Fúngicas
/
Genes Reporter
/
Técnicas do Sistema de Duplo-Híbrido
Tipo de estudo:
Diagnostic_studies
/
Evaluation_studies
Idioma:
En
Ano de publicação:
2004
Tipo de documento:
Article