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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Yeast 9 (1993), S. 723-732 
    ISSN: 0749-503X
    Keywords: Optical trapping ; yeast ; isolation method ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: Individual yeast cells can be successfully isolated and recultured on plates with a new isolation method making use of optical trapping with infrared laser light. The cells can be selected on morphological criteria by high resolution microscopy. The isolation device is constructed from two coverslips separated by spacers, in which selected cells are transferred to a plastic capillary, using the optical trap. To test the procedure, selection experiments were done with a mixture of two Saccharomyces cerevisiae strains, distinguishable both in fluorescence microscopy and on agar plates. These experiments showed that only selected cells were isolated, and close to 100% of the isolated stationary-phase cells formed colonies on agar plates, indicating a high recovery. A lower recovery was obtained with exponential-phase cells, possibly because of a higher sensitivity to laser irradiation. Applications for this method may include the isolation of mutants with altered morphology and the isolation of subpopulations of yeast cultures, for their separate investigation or for the initiation of pure cultures.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 0749-503X
    Keywords: Saccharomyces cerevisiae ; vacuole biogenesis ; phenotypic lag ; optical trapping ; Life and Medical Sciences ; Genetics
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology
    Notes: The conditional vacuolar segregation mutant vac2-1 [Shaw and Wickner (1991) EMBO J. 10,1741-1748] shifted to non-permissive temperature (37°C), forms large-budded cells without a vacuole in the bud, and daughter cells without an apparent vacuole. Some cells still contain normal segregation structures. Structural and biochemical quantification of the segregation defect showed that (i) about 10% of the full-grown buds did not contain a vacuole, (ii) about 15% of the small cells washed out of a population growing in an elutriation chamber at 37°C, did not contain a visible vacuole, and (iii) 15% of the cells per generation lost carboxypeptidase Y activity after proteinase A depletion. Thus, 10-15% of the daughter cells did not inherit vacuolar structures or vacuolar proteolytic activity from the mother cell. To investigate the fate of vacuole-less daughters, these cells were isolated by optical trapping. The isolated cells formed colonies on agar plates that consisted of cells with normal vacuoles, both at 23 and 37°C. Thus, the vacuole-less cells that failed to inherit proteolytic activities from the mother cell apparently give rise to progeny containing structurally normal vacuoles. Time-lapse experiments showed that vacuole-less daughter cells formed vacuolar vesicles that fused into a new vacuole within 30 min. Although new buds only emerged after a vacuole had formed in the mother cell, the temporary lack of a vacuole had little effect on growth rate. The results suggest that an alternative pathway for vacuole formation exists, and that yeast cells may require a vacuole of some minimal size to initiate a new round of budding. © 1997 by John Wiley & Sons, Ltd.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
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