CRISPR-Cas9, the new kid on the block of fungal molecular biology

<span class="paragraphSection"><div class="boxTitle">Abstract</div>Research on fungal pathogens with the aim to identify virulence determinants strictly relies on the generation of defined, recombinant strains, a task that is executed by means of a sophisticated molecular biology toolbox. Recent developments in fungal genome engineering have opened a new frontier by implementing the CRISPR-Cas9 technology, based on expression of the Cas9 endonuclease that is loaded by a single guiding RNA (sgRNA) molecule to target a defined site in the recipient genome. This novel approach has been adapted successfully to engineer fungal genomes, among them the one of the human-pathogenic mould <span style="font-style:italic;">Aspergillus fumigatus</span>. Implementation of the required components was achieved by various means that differ with respect to expression of the Cas9 enzyme and sgRNA delivery. Validation of CRISPR-Cas9-mediated mutagenesis could be executed by targeting selected candidate genes of <span style="font-style:italic;">A. fumigatus</span> to provide a promising perspective for screening and multiplexing approaches to scrutinize the virulome of this opportunistic fungal pathogen in a comprehensive manner, such as by analyzing genetic polymorphisms or the function of gene families.</span>

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