A method for producing a protein of interest by culturing a recombinant eukaryotic cell line comprising an expression construct comprising an adjustable promoter and a nucleic acid molecule encoding a poi under the transcriptional control of such a promoter.

摘要

A method of producing a promoter variant which is a variant of a promoter selected from the group consisting of pG1 consisting of the nucleotide sequence of SEQ ID 1, pG3 consisting of the nucleotide sequence of SEQ ID 2, pG4 consisting of the nucleotide sequence of SEQ ID 4, pG6 consisting of the nucleotide sequence of SEQ ID 3, pG7 consisting of the nucleotide sequence of SEQ ID 5, and pG8 consisting of the nucleotide sequence of SEQ ID 6, which method comprisesa) mutagenesis of the respective promoter nucleotide sequence thereby producing variants comprising a nucleotide sequence of at least 200 bp and having at least 80% sequence identity to the respective promoter nucleotide sequence,b) determining the transcription rate of the variants, andc) selecting a promoter variant from said variants which is capable of producing a protein of interest (POI) in a recombinant eukaryotic cell by expressing a nucleotide sequence encoding said POI under the transcriptional control of said promoter variant, at a transcription rate of at least 20% as compared to the pGAP promoter (SEQ ID NO:13).