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Proteins, Expression, Isolation and Analysis | … The mycelium was collected by filtration and used to extract intracellular proteins after grinding in liquid nitrogen. Fusion protein production was examined by SDS–polyacrylamide gel electrophoresis and western blots using an anti-GFP antibody (Genscript, Nanjing, China) … | Get A Quote |
Filamentous fungi have wide applications in biotechnology. The CRISPR/Cas9 system is a powerful genome-editing method that facilitates genetic alterations of genomes in a variety of organisms. However, a genome-editing approach has not been reported in filamentous fungi. Here, we demonstrated the establishment of a CRISPR/Cas9 system in the filamentous fungus Trichoderma reesei by specific codon optimization and in vitro RNA transcription. It was shown that the CRISPR/Cas9 system was controllable and conditional through inducible Cas9 expression. This system generated site-specific mutations in target genes through efficient homologous recombination, even using short homology arms. This system also provid... More