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Vectors for gene expression and amplification in the yeast Yarrowia lipolytica
Author(s) -
Juretzek Thomas,
Le Dall MarieThérèse,
Mauersberger Stephan,
Gaillardin Claude,
Barth Gerold,
Nicaud JeanMarc
Publication year - 2001
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/1097-0061(20010130)18:2<97::aid-yea652>3.0.co;2-u
Subject(s) - biology , plasmid , gene , promoter , reporter gene , microbiology and biotechnology , genetics , low copy number , expression vector , shuttle vector , escherichia coli , gene expression , vector (molecular biology) , recombinant dna
New vector systems were developed for gene expression in Y. lipolytica . These plasmids contain: (a) as integration target sequences, either a rDNA region or the long terminal repeat zeta of the Y. lipolytica retrotransposon Ylt1 ; (b) the YlURA3 gene as selection marker for Y. lipolytica , either as the non‐defective ura3d1 allele for single integration or the promotor truncated ura3d4 allele for multiple integration; (c) the inducible ICL1 or XPR2 promoters for gene expression; and (d) unique restriction sites for gene insertion. Multiple plasmid integration occurred as inserted tandem‐repeats, which are present at 3–39 copies per cell. A correlation between gene copy number and the expressed enzyme activity was demonstrated with Escherichia coli lacZ as reporter gene under the control of the regulated ICL1 promoter. Increases in copy numbers from 5 to 13 for the lacZ expression cassettes resulted in an up to 10–11‐fold linear increase of the β‐galactosidase activity in multicopy transformants during their growth on ethanol or glucose, compared with the low‐copy replicative plasmid transformants (1.6 plasmid copies). These new tools will enhance the interest in Y. lipolytica as an alternative host for heterologous protein production. Copyright © 2000 John Wiley & Sons, Ltd.