Optimization of yeast cell cycle analysis and morphological characterization by multispectral imaging flow cytometry

Budding yeast Saccharoymyces cerevisiae is a powerful model system for analyzing eukaryotic cell cycle regulation. Yeast cell cycle analysis is typically performed by visual analysis or flow cytometry, and both have limitations in the scope and accuracy of data obtained. This study demonstrates how...

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Bibliographic Details
Published in:Cytometry. Part A 2008-09, Vol.73A (9), p.825-833
Main Authors: Calvert, Meredith E.K, Lannigan, Joanne A, Pemberton, Lucy F
Format: Article
Language:English
Subjects:
budding yeast
Cell Cycle
Cell Cycle Proteins - metabolism
Cytoskeletal Proteins - metabolism
Flow Cytometry - methods
Image Cytometry - methods
MIFC
multipectral imaging flow cytometry
Nap1
Nuclear Proteins - metabolism
Saccharomyces cerevisiae - cytology
Saccharomyces cerevisiae - growth & development
Saccharomyces cerevisiae - metabolism
Saccharomyces cerevisiae Proteins - metabolism
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Summary:Budding yeast Saccharoymyces cerevisiae is a powerful model system for analyzing eukaryotic cell cycle regulation. Yeast cell cycle analysis is typically performed by visual analysis or flow cytometry, and both have limitations in the scope and accuracy of data obtained. This study demonstrates how multispectral imaging flow cytometry (MIFC) provides precise quantitation of cell cycle distribution and morphological phenotypes of yeast cells in flow. Cell cycle analysis of wild-type yeast, nap1Δ, and yeast overexpressing NAP1, was performed visually, by flow cytometry and by MIFC. Quantitative morphological analysis employed measurements of cellular length, thickness, and aspect ratio in an algorithm to calculate a novel feature, bud length. MIFC demonstrated reliable quantification of the yeast cell cycle compared to morphological and flow cytometric analyses. By employing this technique, we observed both the G2/M delay and elongated buds previously described in the nap1Δ strain. Using MIFC, we demonstrate that overexpression of NAP1 causes elongated buds yet only a minor disruption in the cell cycle. The different effects of NAP1 expression level on cell cycle and morphology suggests that these phenotypes are independent. Unlike conventional yeast flow cytometry, MIFC generates complete cell cycle profiles and concurrently offers multiple parameters for morphological analysis. © 2008 International Society for Advancement of Cytometry
ISSN:1552-4922
1552-4930
DOI:10.1002/cyto.a.20609