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Related Experiment Videos

Quantitative analysis of yeast gene function using competition experiments in continuous culture

F Baganz1, A Hayes, R Farquhar

  • 1Department of Biomolecular Sciences, UMIST, Manchester, U.K.

Yeast (Chichester, England)
|December 16, 1998
PubMed
Summary

This study introduces a quantitative method using competition experiments in chemostat cultures to assess gene function by measuring the growth rate effects of gene deletions in Saccharomyces cerevisiae (S. cerevisiae). The findings demonstrate the suitability of this approach for distinguishing between mutants with similar phenotypes.

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Area of Science:

  • * Molecular and Systems Biology
  • * Quantitative Biology
  • * Yeast Genetics

Background:

  • * Evaluating gene function is crucial for understanding cellular processes.
  • * Metabolic Control Analysis (MCA) offers a quantitative framework for gene function evaluation.
  • * Assessing the impact of gene deletions on growth rate (fitness) in Saccharomyces cerevisiae (S. cerevisiae) is a key initial step.

Purpose of the Study:

  • * To establish a quantitative method for determining the fitness effects of gene deletions in S. cerevisiae.
  • * To validate the use of competition experiments in chemostat cultures for this purpose.
  • * To assess the accuracy and reproducibility of quantitative PCR (qPCR) based methods for measuring strain proportions.

Main Methods:

  • * Competition experiments were conducted in chemostat cultures using deletion mutants and a standard S. cerevisiae strain.

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  • * Quantitative PCR (qPCR) was employed to measure the relative proportions of competing strains.
  • * Densitometry and GeneScan analysis were compared for their accuracy in determining strain proportions (10-90% of the population).

    • Main Results:

    • * Both densitometry and GeneScan analysis demonstrated high accuracy and reproducibility in quantifying strain proportions.
    • * Competition experiments in chemostat cultures successfully distinguished between deletion mutants with qualitatively similar phenotypes.
    • * The method proved effective under six different physiological conditions.

    Conclusions:

    • * Competition experiments in continuous culture provide a robust quantitative method for assessing gene deletion effects on S. cerevisiae fitness.
    • * This approach is valuable for distinguishing subtle fitness differences between mutants.
    • * The validated qPCR-based method enables precise quantification of strain proportions crucial for genetic analysis.