ISSN:
1432-0983
Schlagwort(e):
Schizosaccharomyces pombe
;
Saccharomyces cerevisiae
;
β-glucuronidase
;
Colony colour assay
;
Fluorometric assay
Quelle:
Springer Online Journal Archives 1860-2000
Thema:
Biologie
Notizen:
Summary Expression of the β-galactosidase gene in yeast has served as a screening marker for many purposes. Here it is shown that in two yeasts, Saccharomyces cerevisiae and Schizosaccharomyces pombe, the β-glucuronidase (GUS) gene can be used as an alternative marker. Since the histochemical substrate can not be taken up by yeast cells, direct colony screening of plates was found to be impossible. However, by a replica plating technique, GUS expression became visibly detectable within 10 min when the GUS gene was strongly expressed. The staining method could still be performed for expression at a 100-fold lower level, but incubation times of several hours were needed. Furthermore, specific GUS expression levels of yeast protein extracts could be quantified by a fluorometric assay which is both very simple to perform and highly sensitive. Since the GUS gene can also tolerate large N-terminal fusions, this method should be particularly attractive for studying such diverse problems as transcriptional and translational regulation or subcellular localization in yeast.
Materialart:
Digitale Medien
URL:
http://dx.doi.org/10.1007/BF00317075
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