A large-scale sonication assay for cell wall mutant analysis in yeast.
The complete yeast genome contains a large number of genes of unknown biological function. Simple, rapid and reliable specific screens are valuable tools in exploring gene function via systematic phenotypic analysis of large mutant collections. This report provides a new approach for monitoring changes in cell wall strength, based on the deleterious effects caused by ultrasound on the yeast cell surface. Sonication can thus be used for the screening of mutants affected in the architecture or stability of the cell wall, since such mutants are expected to have an altered sensitivity to this treatment compared to that of a wild-type. The experimental procedure, consisting in the quantification of damaged cells after a mild sonication treatment, by means of flow cytometry, can be applied on a large scale. The usefulness of the sonication assay as a primary screen for cell wall-related mutants is evaluated on the collection of calcofluor white-hypersensitive and -resistant mutants obtained by Lussier et al. (1997). A further phenotypic characterization of the sonication-hypersensitive mutants within the calcofluor white collection is also presented.
Departamento de Microbiología II, Facultad de Farmacia, Universidad Complutense de Madrid, Madrid, Spain.
C
C
Ruiz
V J
VJ
Cid
M
M
Lussier
M
M
Molina
C
C
Nombela
D001557:Benzenesulfonates / Q000494:pharmacology; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology; D002473:Cell Wall / Q000737:chemistry / Q000502:physiology*; D005434:Flow Cytometry; D005800:Genes, Fungal; D009154:Mutation; D010641:Phenotype; D013010:Sonication; D015003:Yeasts / Q000187:drug effects / Q000235:genetics* / Q000648:ultrastructure
D001557:Benzenesulfonates; D002110:Caffeine; C007061:C.I. Fluorescent Brightening Agent 28; D002118:Calcium