ArticleBMC biotechnology2019
Fast gene disruption in Trichoderma reesei using in vitro assembled Cas9/gRNA complex.
Article in BMC biotechnology, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers, 1 of them a synthesis that pooled it.
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Who cites it
24 citing papers in PubMed, 1 synthesis or guideline pooled it.
- BioprospectingFrontiers in fungal biology · 2021Pooled it
- Optimization of the Genome Editing CRISPR-Cas9 Technology in Scedosporium apiospermum.Mycopathologia · 2025Article
- Development of a CRISPR/Cas9 RNP-mediated genetic engineering system in Paecilomyces variotii.Journal of microbiology (Seoul, Korea) · 2025Article
- Simply cut out - Combining CRISPR/Cas9 RNPs and transiently selected telomere vectors for marker free-gene deletion inFrontiers in genome editing · 2025Article
- CRISPR-Based Gene Editing: a Modern Approach for Study and Treatment of Cancer.Applied biochemistry and biotechnology · 2024Review
- Edible mycelium bioengineered for enhanced nutritional value and sensory appeal using a modular synthetic biology toolkit.Nature communications · 2024Article
- An efficient CRISPR/Cas9 genome editing system based on a multiple sgRNA processing platform in Trichoderma reesei for strain improvement and enzyme production.Biotechnology for biofuels and bioproducts · 2024Article
- Article
- Current genetic strategies to investigate gene functions in Trichoderma reesei.Microbial cell factories · 2023Review
- A Simple CRISPR/Cas9 System for Efficiently Targeting Genes of Aspergillus SectionMicrobiology spectrum · 2023Article
- CRISPR-Cas12a induced DNA double-strand breaks are repaired by multiple pathways with different mutation profiles in Magnaporthe oryzae.Nature communications · 2022Article
- The contribution of DNA repair pathways to genome editing and evolution in filamentous pathogens.FEMS microbiology reviews · 2022Review
- Use of CRISPR-Cas tools to engineer Trichoderma species.Microbial biotechnology · 2022Review
- CRISPR/Cas9-Based Genome Editing and Its Application inJournal of fungi (Basel, Switzerland) · 2022Review
- Review
- Efficient genome editing in filamentous fungi via an improved CRISPR-Cas9 ribonucleoprotein method facilitated by chemical reagents.Microbial biotechnology · 2021Article
- An overview on current molecular tools for heterologous gene expression in Trichoderma.Fungal biology and biotechnology · 2021Review
- CRISPR/Cas9-mediated genome editing in Penicillium oxalicum and Trichoderma reesei using 5S rRNA promoter-driven guide RNAs.Biotechnology letters · 2021Article
- Generation of Trichoderma harzianum with pyr4 auxotrophic marker by using the CRISPR/Cas9 system.Scientific reports · 2021Article
- Applications of CRISPR/Cas9 in the Synthesis of Secondary Metabolites in Filamentous Fungi.Frontiers in microbiology · 2021Review
Corrections and comments
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2 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundCRISPR/Cas9 has wide application potentials in a variety of biological species including Trichoderma reesei, a filamentous fungus workhorse for cellulase production. However, expression of Cas9 heterologously in the host cell could be time-consuming and sometimes even troublesome.
resultsWe tested two gene disruption methods in T. reesei using CRISPR/Cas9 in this study. The intracellularly expressed Cas9 led to unexpected off-target gene disruption in T. reesei QM9414, favoring inserting 9- or 12-bp at 70- and 100-bp downstream of the targeted ura5. An alternative method was, therefore, established by assembling Cas9 and gRNA in vitro, followed by transformation of the ribonucleoprotein complex with a plasmid containing the pyr4 marker gene into T. reesei TU-6. When the gRNA targeting cbh1 was used, eight among the twenty seven transformants were found to lose the ability to express CBH1, indicative of successful cbh1 disruption through genome editing. Large DNA fragments including the co-transformed plasmid, chromosomal genes, or a mixture of these nucleotides, were inserted in the disrupted cbh1 locus.
conclusionsDirect transformation of Cas9/gRNA complex into the cell is a fast means to disrupt a gene in T. reesei and may find wide applications in strain improvement and functional genomics study.
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