Genomic deletions in Aureobasidium pullulans by an AMA1 plasmid for gRNA and CRISPR/Cas9 expression.

Autor: Masi A; Christian Doppler Laboratory for Optimised Expression of Carbohydrate-Active Enzymes, Institute of Chemical, Environmental and Bioscience Engineering, TU Wien, Gumpendorfer Str. 1a, A-1060, Vienna, Austria.; Institute of Chemical, Environmental and Bioscience Engineering, TU Wien, Gumpendorfer Str. 1a, A-1060, Vienna, Austria., Wögerbauer K; Institute of Chemical, Environmental and Bioscience Engineering, TU Wien, Gumpendorfer Str. 1a, A-1060, Vienna, Austria., Mach RL; Institute of Chemical, Environmental and Bioscience Engineering, TU Wien, Gumpendorfer Str. 1a, A-1060, Vienna, Austria., Mach-Aigner AR; Christian Doppler Laboratory for Optimised Expression of Carbohydrate-Active Enzymes, Institute of Chemical, Environmental and Bioscience Engineering, TU Wien, Gumpendorfer Str. 1a, A-1060, Vienna, Austria. astrid.mach-aigner@tuwien.ac.at.; Institute of Chemical, Environmental and Bioscience Engineering, TU Wien, Gumpendorfer Str. 1a, A-1060, Vienna, Austria. astrid.mach-aigner@tuwien.ac.at.
Jazyk: angličtina
Zdroj: Fungal biology and biotechnology [Fungal Biol Biotechnol] 2024 Jun 01; Vol. 11 (1), pp. 6. Date of Electronic Publication: 2024 Jun 01.
DOI: 10.1186/s40694-024-00175-4
Abstrakt: Background: Aureobasidium pullulans is a generalist polyextremotolerant black yeast fungus. It tolerates temperatures below 0 °C or salt concentrations up to 18%, among other stresses. A. pullulans genome sequencing revealed a high potential for producing bioactive metabolites. Only few molecular tools exist to edit the genome of A. pullulans, hence it is important to make full use of its potential. Two CRISPR/Cas9 methods have been proposed for the protoplast-based transformation of A. pullulans. These methods require the integration of a marker gene into the locus of the gene to be deleted, when the deletion of this gene does not yield a selectable phenotype. We present the adaptation of a plasmid-based CRISPR/Cas9 system developed in Aspergillus niger for A. pullulans to create deletion strains.
Results: The A. niger CRISPR/Cas9 plasmid led to efficient genomic deletions in A. pullulans. In this study, strains with deletions ranging from 30 to 862 bp were obtained by using an AMA1 plasmid-based genome editing strategy.
Conclusion: The CRISPR/Cas9 transformation system presented in this study provides new opportunities for strain engineering of A. pullulans. This system allows expression of Cas9 and antibiotic resistance while being easy to adapt. This strategy could open the path to intensive genomic engineering in A. pullulans.
(© 2024. The Author(s).)
Databáze: MEDLINE
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