ArticleScientific reports2016
Harnessing heterologous and endogenous CRISPR-Cas machineries for efficient markerless genome editing in Clostridium.
Article in Scientific reports, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 91 papers.
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Who cites it
91 citing papers in PubMed, 179 citations in OpenAlex.
- Review
- Sensitizing carbapenem-resistant Klebsiella pneumoniae using a compact Cascade-Cas3 system.International microbiology : the official journal of the Spanish Society for Microbiology · 2026Article
- Transferable genetic toolsets for nonmodel gut Clostridia enable in vivo reversible control of metabolite production.Nature biotechnology · 2026Article
- Target discrimination and PAM profiling of the Thermotoga maritima type I-B CRISPR system.The Biochemical journal · 2026Article
- Functional characterization of a type I-F1 CRISPR-cas system from the clinical isolate Shewanella xiamenensis Sh95 reveals constitutive activity and plasmid-curing capability.Scientific reports · 2026Article
- Sequential structural rearrangements at the PAM-distal site of a type I-F3 CRISPR-Cas effector enabling RNA-guided DNA transposition.Nucleic acids research · 2026Article
- Exploiting the Endogenous Type II-A CRISPR-Cas System for Functional Engineering of Probiotic Lacticaseibacillus rhamnosus GG.Microbial biotechnology · 2026Article
- Hi-TARGET: a fast, efficient and versatile CRISPR type I-B genome editing tool for the thermophilic acetogen Thermoanaerobacter kivui.Biotechnology for biofuels and bioproducts · 2025Article
- Deletion of atypical type II restriction genes in Clostridium cellulovorans using a Cas9-based gene editing system.Applied microbiology and biotechnology · 2025Article
- Emerging trends in genome integration tools for precision engineering of diverse bacterial species.Synthetic biology (Oxford, England) · 2025Review
- A Consecutive Genome Engineering Method Reveals a New Phenotype and Regulation of Glucose and Glycerol Utilization inEngineering in life sciences · 2025Article
- Research Progress on the Mechanism and Application of the Type I CRISPR-Cas System.International journal of molecular sciences · 2024Review
- Review
- CRISPR-Cas9-based genome-editing technologies in engineering bacteria for the production of plant-derived terpenoids.Engineering microbiology · 2024Review
- Structure and genome editing of type I-B CRISPR-Cas.Nature communications · 2024Article
- A type I-F CRISPRi system unveils the novel role of CzcR in modulating multidrug resistance ofMicrobiology spectrum · 2023Article
- Dissecting the Arginine and Lysine Biosynthetic Pathways and Their Relationship in Haloarchaeon Natrinema gari J7-2 via Endogenous CRISPR-Cas System-Based Genome Editing.Microbiology spectrum · 2023Article
- Repurposing the atypical type I-G CRISPR system for bacterial genome engineering.Microbiology (Reading, England) · 2023Article
- CRISPR-Based Gene Editing inPharmaceuticals (Basel, Switzerland) · 2023Review
- Pangenomic Study of Fusobacterium nucleatum Reveals the Distribution of Pathogenic Genes and Functional Clusters at the Subspecies and Strain Levels.Microbiology spectrum · 2023Article
31 more citing papers are in PubMed but not listed here.
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Authors and funding
5 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Application of CRISPR-Cas9 systems has revolutionized genome editing across all domains of life. Here we report implementation of the heterologous Type II CRISPR-Cas9 system in Clostridium pasteurianum for markerless genome editing. Since 74% of species harbor CRISPR-Cas loci in Clostridium, we also explored the prospect of co-opting host-encoded CRISPR-Cas machinery for genome editing. Motivation for this work was bolstered from the observation that plasmids expressing heterologous cas9 result in poor transformation of Clostridium. To address this barrier and establish proof-of-concept, we focus on characterization and exploitation of the C. pasteurianum Type I-B CRISPR-Cas system. In silico spacer analysis and in vivo interference assays revealed three protospacer adjacent motif (PAM) sequences required for site-specific nucleolytic attack. Introduction of a synthetic CRISPR array and cpaAIR gene deletion template yielded an editing efficiency of 100%. In contrast, the heterologous Type II CRISPR-Cas9 system generated only 25% of the total yield of edited cells, suggesting that native machinery provides a superior foundation for genome editing by precluding expression of cas9 in trans. To broaden our approach, we also identified putative PAM sequences in three key species of Clostridium. This is the first report of genome editing through harnessing native CRISPR-Cas machinery in Clostridium.
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