Functional Reconstitution of a Fungal Natural Product Gene Cluster by Advanced Genome Editing.
Filamentous fungi produce varieties of natural products even in a strain dependent manner. However, the genetic basis of chemical speciation between strains is still widely unknown. One example is trypacidin, a natural product of the opportunistic human pathogen Aspergillus fumigatus, which is not produced among different isolates. Combining computational analysis with targeted gene editing, we could link a single nucleotide insertion in the polyketide synthase of the trypacidin biosynthetic pathway and reconstitute its production in a nonproducing strain. Thus, we present a CRISPR/Cas9-based tool for advanced molecular genetic studies in filamentous fungi, exploiting selectable markers separated from the edited locus.
Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute (HKI) , 07745 Jena, Germany.|Institute of Microbiology, Friedrich Schiller University Jena , 07743 Jena, Germany.
Jakob
J
Weber
Leibniz Research Group - Biobricks of Microbial Natural Product Syntheses, Leibniz Institute for Natural Product Research and Infection Biology (HKI) , 07745 Jena, Germany.
Vito
V
Valiante
Eukaryotic Molecular Cell Biology, Section for Eukaryotic Biotechnology, Department of Systems Biology, Technical University of Denmark , Søltofts Plads, 2800 Kongens Lyngby, Denmark.
Christina S
CS
Nødvig
Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute (HKI) , 07745 Jena, Germany.|Institute of Microbiology, Friedrich Schiller University Jena , 07743 Jena, Germany.
Derek J
DJ
Mattern
Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute (HKI) , 07745 Jena, Germany.
Rebecca A
RA
Slotkowski
Eukaryotic Molecular Cell Biology, Section for Eukaryotic Biotechnology, Department of Systems Biology, Technical University of Denmark , Søltofts Plads, 2800 Kongens Lyngby, Denmark.
Uffe H
UH
Mortensen
Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute (HKI) , 07745 Jena, Germany.|Institute of Microbiology, Friedrich Schiller University Jena , 07743 Jena, Germany.
Axel A
AA
Brakhage
D001232:Aspergillus fumigatus / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity; D001688:Biological Products / Q000378:metabolism; D064113:CRISPR-Cas Systems; D000072669:Gene Editing / Q000379:methods*; D005800:Genes, Fungal; D006801:Humans; D005810:Multigene Family; D009183:Mycotoxins / Q000096:biosynthesis / Q000235:genetics; D048630:Polyketide Synthases / Q000235:genetics / Q000378:metabolism; D058615:Synthetic Biology
D001688:Biological Products; D009183:Mycotoxins; D048630:Polyketide Synthases
Aspergillus fumigatus; CRISPR/Cas9; gene-editing; split-marker; trypacidin