Flow cytometry analysis of recombinant Saccharomyces cerevisiae populations

Cytometry. 1986 Mar;7(2):132-41. doi: 10.1002/cyto.990070203.

Abstract

A new fluorescent stain has been developed for detecting cloned beta-galactosidase activity in individual cells of Saccharomyces cerevisiae by flow cytometry. The staining reaction is based on enzymatic cleavage of alpha-naphthol-beta-D-galactopyranoside by intracellular beta-galactosidase and trapping of the liberated naphthol by hexazoniumpararosaniline yielding a fluorescent, insoluble end product. This stain, in connection with an appropriate host strain, has been applied for detecting plasmids encoding inducible beta-galactosidase in unstable recombinant cell populations carrying plasmids with different origins of replication. The method enables rapid determination of the fraction of plasmid-containing cells as well as quantitation of intracellular beta-galactosidase content by kinetic enzyme assay. Inducibility of the marker enzyme is important for maintaining correlation between enzyme and gene content.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Flow Cytometry*
  • Fluorescent Dyes
  • Galactosidases / analysis*
  • Kinetics
  • Plasmids
  • Recombination, Genetic
  • Saccharomyces cerevisiae / enzymology*
  • Saccharomyces cerevisiae / genetics
  • Staining and Labeling
  • beta-Galactosidase / analysis*

Substances

  • Fluorescent Dyes
  • Galactosidases
  • beta-Galactosidase