Functional reconstitution of the Trypacidin Gene Cluster in Aspergillus fumigatus by Advanced Gene Editing

J. Webera,b, V. Valiantec, C.S. Nødvigd, D.J. Matterna,b, R.A. Slotkowskia,b, U.H. Mortensend and A.A. Brakhagea,b

Author address: 

aDepartment of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology – Hans Knöll Institute (HKI) –, Jena, Germany bInstitute of Microbiology, Friedrich Schiller University Jena, Germany cLeibniz Research Group – Biobricks of Microbial Natural Product Syntheses, Leibniz Institute for Natural Product Research and Infection Biology (HKI), Jena, Germany dEukaryotic Molecular Cell Biology, Section for Eukaryotic Biotechnology, Department of Systems Biology, Technical University of Denmark, Søltofts Plads, Kongens Lyngby, Denmark

Abstract: 

The human pathogenic fungus Aspergillus fumigatus is known to produce various spore-borne natural products. One of these is the polyketide trypacidin which has been shown to be involved in the interactions with alveolar macrophages as well as with the amoeba Dictyostelium discoideum. Even though recent studies could elucidate the corresponding gene cluster in A. fumigatus, it still remained elusive why several isolates do not produce trypacidin. We addressed this question employing a CRISPR/Cas9-based gene editing strategy. Thus, we could link a single nucleotide insertion in the polyketide synthase of the trypacidin biosynthetic pathway and also reconstitute its production in a nonproducing strain. In addition, we developed a split-marker approach for the selection of edited strains, since the selectable marker could not be directly linked to the target site. The here established tool could be useful in next generation fungal genetics e.g. for the investigation of single nucleotide polymorphism, or amino acid substitutions.

2017

abstract No: 

77

Full conference title: 

The Fourteenth International Aspergillus Meeting, Asilomar Conference Center, Pacific Grove, CA, USA
    • Asperfest 14 (2017)