Flow cytometry analysis of recombinant Saccharomyces cerevisiae populations

Friedrich Srienc, Judith L. Campbell, James E. Bailey

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

A new fluorescent stain has been developed for detecting cloned β‐galactosidase activity in individual cells of Saccharomyces cerevisiae by flow cytometry. The staining reaction is based on enzymatic cleavage of α‐naphthol‐β‐D‐galactopyranoside by intracellular β‐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 β‐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 β‐galactosidase content by kinetic enzyme assay. Inducibility of the marker enzyme is important for maintaining correlation between enzyme and gene content.

Original languageEnglish (US)
Pages (from-to)132-141
Number of pages10
JournalCytometry
Volume7
Issue number2
DOIs
StatePublished - Mar 1986

Keywords

  • Saccharomyces cerevisiae
  • flow cytometry
  • plasmid stability
  • β‐galactosidase stain

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