title: CRISPR/Cas9-mediated efficient genome editing via blastospore-based transformation in entomopathogenic fungus Beauveria bassiana.
abstract: Beauveria bassiana is an environmentally friendly alternative to chemical insecticides against various agricultural insect pests and vectors of human diseases. However, its application has been limited due to slow kill and sensitivity to abiotic stresses. Understanding of the molecular pathogenesis and physiological characteristics would facilitate improvement of the fungal performance. Loss-of-function mutagenesis is the most powerful tool to characterize gene functions, but it is hampered by the low rate of homologous recombination and the limited availability of selectable markers. Here, by combining the use of uridine auxotrophy as recipient and donor DNAs harboring auxotrophic complementation gene ura5 as a selectable marker with the blastospore-based transformation system, we established a highly efficient, low false-positive background and cost-effective CRISPR/Cas9-mediated gene editing system in B. bassiana. This system has been demonstrated as a simple and powerful tool for targeted gene knock-out and/or knock-in in B. bassiana in a single gene disruption. We further demonstrated that our system allows simultaneous disruption of multiple genes via homology-directed repair in a single transformation. This technology will allow us to study functionally redundant genes and holds significant potential to greatly accelerate functional genomics studies of B. bassiana.
authors.affiliation: CAS Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.|University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Jingjing
authors.initials: J
authors.lastname: Chen
authors.affiliation: CAS Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Yiling
authors.initials: Y
authors.lastname: Lai
authors.affiliation: CAS Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.|University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Wang
authors.affiliation: CAS Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Suzhen
authors.initials: S
authors.lastname: Zhai
authors.affiliation: CAS Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Gen
authors.initials: G
authors.lastname: Zou
authors.affiliation: CAS Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Zhihua
authors.initials: Z
authors.lastname: Zhou
authors.affiliation: CAS Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.|University of Chinese Academy of Sciences, Beijing 100049, China.
authors.forename: Chunlai
authors.initials: C
authors.lastname: Cui
authors.affiliation: CAS Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
authors.forename: Sibao
authors.initials: S
authors.lastname: Wang
mesh_terms: D052982:Beauveria / Q000235:genetics* / Q000254:growth & development; D064113:CRISPR-Cas Systems; D000072669:Gene Editing; D018390:Gene Targeting / Q000379:methods*; D005800:Genes, Fungal; D005822:Genetic Vectors; D006801:Humans; D011995:Recombination, Genetic; D014170:Transformation, Genetic; D051981:Uracil-DNA Glycosidase / Q000037:antagonists & inhibitors* / Q000235:genetics; D014529:Uridine / Q000378:metabolism*
chemical_list: D051981:Uracil-DNA Glycosidase; D014529:Uridine