Application of CRISPR/Cas9 Tools for Genome Editing in the White-Rot Fungus Dichomitus squalens

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Kowalczyk , J E , Saha , S & Mäkelä , M R 2021 , ' Application of CRISPR/Cas9 Tools for Genome Editing in the White-Rot Fungus Dichomitus squalens ' , Biomolecules , vol. 11 , no. 10 , 1526 . https://doi.org/10.3390/biom11101526

Title: Application of CRISPR/Cas9 Tools for Genome Editing in the White-Rot Fungus Dichomitus squalens
Author: Kowalczyk, Joanna E.; Saha, Shreya; Mäkelä, Miia R.
Other contributor: University of Helsinki, Department of Microbiology
University of Helsinki, Plant-Virus Interactions
University of Helsinki, Helsinki Institute of Sustainability Science (HELSUS)




Date: 2021-10
Language: eng
Number of pages: 15
Belongs to series: Biomolecules
ISSN: 2218-273X
DOI: https://doi.org/10.3390/biom11101526
URI: http://hdl.handle.net/10138/336689
Abstract: Dichomitus squalens is an emerging reference species that can be used to investigate white-rot fungal plant biomass degradation, as it has flexible physiology to utilize different types of biomass as sources of carbon and energy. Recent comparative (post-) genomic studies on D. squalens resulted in an increasingly detailed knowledge of the genes and enzymes involved in the lignocellulose breakdown in this fungus and showed a complex transcriptional response in the presence of lignocellulose-derived compounds. To fully utilize this increasing amount of data, efficient and reliable genetic manipulation tools are needed, e.g., to characterize the function of certain proteins in vivo and facilitate the construction of strains with enhanced lignocellulolytic capabilities. However, precise genome alterations are often very difficult in wild-type basidiomycetes partially due to extremely low frequencies of homology directed recombination (HDR) and limited availability of selectable markers. To overcome these obstacles, we assessed various Cas9-single guide RNA (sgRNA) ribonucleoprotein (RNP) -based strategies for selectable homology and non-homologous end joining (NHEJ) -based gene editing in D. squalens. We also showed an induction of HDR-based genetic modifications by using single-stranded oligodeoxynucleotides (ssODNs) in a basidiomycete fungus for the first time. This paper provides directions for the application of targeted CRISPR/Cas9-based genome editing in D. squalens and other wild-type (basidiomycete) fungi.
Subject: Dichomitus squalens
<p>CRISPR/Cas9 & nbsp;</p>
genome editing
ribonucleoprotein
single-stranded oligonucleotides
GENE REPLACEMENT
MARKER GENE
RESISTANCE
SYSTEM
TRANSFORMATION
MANIPULATION
DEGRADATION
CARBOXIN
BIOMASS
ACID
1184 Genetics, developmental biology, physiology
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