CRISPR/Cas9 Based Genome Editing of Penicillium chrysogenum
Journal article, 2016
CRISPR/Cas9 based systems have emerged as versatile platforms for precision genome editing in a wide range of organisms. Here we have developed powerful CRISPR/Cas9 tools for marker-based and marker-free genome modifications in Penicillium chrysogenum, a model filamentous fungus and industrially relevant cell factory. The developed CRISPR/Cas9 toolbox is highly flexible and allows editing of new targets with minimal cloning efforts. The Cas9 protein and the sgRNA can be either delivered during transformation, as preassembled CRISPR-Cas9 ribonucleoproteins (RNPs) or expressed from an AMA1 based plasmid within the cell. The direct delivery of the Cas9 protein with in vitro synthesized sgRNA to the cells allows for a transient method for genome engineering that may rapidly be applicable for other filamentous fungi. The expression of Cas9 from an AMA1 based vector was shown to be highly efficient for marker-free gene deletions.
*Penicillium chrysogenum
Guide
Oligonucleotides/genetics
Fungal
*CRISPR/Cas9
Genetic Vectors
Penicillium chrysogenum/*genetics
RNA
*Rnp
Bacterial Proteins/genetics
Genetic Markers
*marker-free gene deletion
Genome
*genome editing
Gene Deletion
DNA Repair
Gene Editing/*methods
Endonucleases/genetics
*CRISPR-Cas Systems
Gene Targeting/methods
Author
C. Pohl
J. A. Kiel
A. J. Driessen
R. A. Bovenberg
Yvonne Nygård
Chalmers, Biology and Biological Engineering, Industrial Biotechnology
ACS Synthetic Biology
2161-5063 (eISSN)
Vol. 5 7 754-64Subject Categories (SSIF 2011)
Other Industrial Biotechnology
DOI
10.1021/acssynbio.6b00082