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Title
Repressible promoters A novel tool to generate conditional mutants in Pichia pastoris
AuthorDelic, Marizela ; Mattanovich, Diethard ; Gasser, Brigitte
Published in
Microbial Cell Factories, 2013, Vol. 12, 6 page
PublishedBioMed Central (BMC), 2013
LanguageEnglish
Document typeJournal Article
Keywords (EN)Repressible promoter / Pichia pastoris / Promoter exchange
ISSN1475-2859
URNurn:nbn:at:at-ubbw:3-1168 Persistent Identifier (URN)
DOIdoi:10.1186/1475-2859-12-6 
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 The work is publicly available
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Abstract (English)

Background:

Repressible promoters are a useful tool for down-regulating the expression of genes, especially those that affect cell viability, in order to study cell physiology. They are also popular in biotechnological processes, like heterologous protein production.

Results:

Here we present five novel repressible Pichia pastoris promoters of different strength: P SER1 , P MET3 , P THR1 , P PIS1 and P THI11 . eGFP was expressed under the control of each of these promoters and its fluorescence could be successfully decreased in liquid culture by adding different supplements. We also expressed the essential genes with different native promoter strength, ERO1 and PDI1, under the control of two of the novel promoters. In our experiments, a clear down-regulation of both repressible promoters on transcriptional level could be achieved. Compared to the transcript levels of these two genes when expressed under the control of their native promoters, only ERO1 was significantly down-regulated.

Conclusion:

Our results show that all of the novel promoters can be used for repression of genes in liquid culture. We also came to the conclusion that the choice of the repressible promoter is of particular importance. For a successful repression experiment it is crucial that the native promoter of a gene and the repressible promoter in its non-repressed state are of similar strength.