Synthetic Core Promoters for <i>Pichia pastoris</i>
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Abstract
Synthetic
promoters are commonly used tools for circuit design
or high level protein production. Promoter engineering efforts in
yeasts, such as <i>Saccharomyces cerevisiae</i> and <i>Pichia pastoris</i> have mostly been focused on altering upstream regulatory
sequences such as transcription factor binding sites. In higher eukaryotes
synthetic core promoters, directly needed for transcription initiation
by RNA Polymerase II, have been successfully designed. Here we report
the first synthetic yeast core promoter for <i>P. pastoris</i>, based on natural yeast core promoters. Furthermore we used this
synthetic core promoter sequence to engineer the core promoter of
the natural <i>AOX1</i> promoter, thereby creating a set
of core promoters providing a range of different expression levels.
As opposed to engineering strategies of the significantly longer entire
promoter, such short core promoters can directly be added on a PCR
primer facilitating library generation and are sufficient to obtain
variable expression yields