Compositions and methods using the yeast YLR110C promoter
AssigneeAstraZeneca AB
InventorsGraham P Beifield, Caroline Oakley
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Abstract
The invention provides novel yeast promoters useful for controlling the expression of homologous and heterologous nucleic acid molecules in yeast cells. The yeast promoters are induced by a fermentable carbon source, such as glucose, or a non-fermentable carbon source, such as ethanol, or both. Therefore, expression of nucleic acid molecules encoding a polypeptide under the control of the novel yeast promoters may be regulated by varying the level of a fermentable carbon source, or a non-fermentable carbon source, or both.
Claims15
An isolated and purified polynucleotide consisting of SEQ ID NO:1 wherein the polynucleotide is operative as a promoter to express a nucleic acid molecule encoding a polypeptide when operably linked to said nucleic acid molecule.
A method for producing a polypeptide comprising the steps of:
- (a) cloning a nucleic acid molecule encoding the polypeptide into an expression vector selected from the group consisting of pYLR110P+luc and pYLR110P, wherein the nucleic acid molecule is operably linked to a promoter of the expression vector;
- (b) transforming a culture of yeast cells with the yeast expression vector;
- (c) maintaining the yeast cells in culture so that the polypeptide is expressed;
- and
- (d) recovering the polypeptide.
A method of identifying a promoter fragment, wherein the fragment has promoter activity comprising the steps of:
- (a) generating a fragment comprising at least 17 contiguous nucleotides of an isolated and purified polynucleotide which is SEQ ID NO:1;
- (b) cloning the fragment into a yeast expression vector, wherein the fragment is operably linked to a reporter gene;
- (c) transforming yeast cells with the yeast expression vector;
- (d) growing the yeast cells in yeast cell culture under conditions favorable for expression of the reporter gene;
- and
- (e) assaying the yeast culture for a reporter protein expressed by the reporter gene;
- wherein expression of the reporter gene indicates the fragment has promoter activity.
Dependencies are inferred from the claim text.