ArticleApplied and environmental microbiology2016
Development of a CRISPR-Cas9 Tool Kit for Comprehensive Engineering of Bacillus subtilis.
Article in Applied and environmental microbiology, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 80 papers.
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Who cites it
80 citing papers in PubMed, 198 citations in OpenAlex.
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- An inducer-independent, single-plasmid CRISPR-Cas9 system for genome editing in Bacillus species.Applied microbiology and biotechnology · 2026Article
- SynBio-DSP: an integrated synthetic biology platform with fed-batch fermentation and continuous downstream processing for sustainable production of thermostable cellulases.Biotechnology for biofuels and bioproducts · 2026Article
- Base Editors for Engineering Industrial Microorganisms: Types, Applications, and Future Perspectives.Biotechnology journal · 2026Review
- Unlocking the Potential for Genetic Engineering of the Straw-Degrading MushroomJournal of fungi (Basel, Switzerland) · 2026Article
- Development of a Tool for High-Efficiency, Markerless and Iterative Genome Editing in Shouchella clausii.Microbial biotechnology · 2026Article
- Advanced applications of synthetic biology technology in biosynthesis of bioactive compounds from medicinal plants.Chinese herbal medicines · 2026Review
- Advances in gene editing tools for four typical Gram-positive bacteria.Frontiers in microbiology · 2026Review
- A three-plasmid-containing CRISPR-Cas9 platform to engineerApplied and environmental microbiology · 2025Article
- LeDNA: a cut-and-build toolkit to democratize CRISPR gene editing technology education.Nature biotechnology · 2025Article
- Chromosomal and Plasmid-Based CRISPRi Platforms for Conditional Gene Silencing inInternational journal of molecular sciences · 2025Article
- CRISPR-based genetic tools for the study of host-microbe interactions.Infection and immunity · 2025Review
- Unlocking Casein Bioactivity: Lactic Acid Bacteria and Molecular Strategies for Peptide Release.International journal of molecular sciences · 2025Review
- Expanding genetic engineering capabilities in Vibrio natriegens with the Vnat Collection.Nucleic acids research · 2025Article
- Exploring the Probiotic Potential of Bacteroides spp. Within One Health Paradigm.Probiotics and antimicrobial proteins · 2025Review
- CRISPR-Cas9-based genome-editing technologies in engineering bacteria for the production of plant-derived terpenoids.Engineering microbiology · 2024Review
- Recent Advances in Metabolic Engineering for the Biosynthesis of Phosphoenol Pyruvate-Oxaloacetate-Pyruvate-Derived Amino Acids.Molecules (Basel, Switzerland) · 2024Review
- Biocomposite thermoplastic polyurethanes containing evolved bacterial spores as living fillers to facilitate polymer disintegration.Nature communications · 2024Article
- Advances in regulating vitamin KWorld journal of microbiology & biotechnology · 2023Review
20 more citing papers are in PubMed but not listed here.
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
unlabelledThe establishment of a clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9 system for strain construction in Bacillus subtilis is essential for its progression toward industrial utility. Here we outline the development of a CRISPR-Cas9 tool kit for comprehensive genetic engineering in B. subtilis In addition to site-specific mutation and gene insertion, our approach enables continuous genome editing and multiplexing and is extended to CRISPR interference (CRISPRi) for transcriptional modulation. Our tool kit employs chromosomal expression of Cas9 and chromosomal transcription of guide RNAs (gRNAs) using a gRNA transcription cassette and counterselectable gRNA delivery vectors. Our design obviates the need for multicopy plasmids, which can be unstable and impede cell viability. Efficiencies of up to 100% and 85% were obtained for single and double gene mutations, respectively. Also, a 2.9-kb hyaluronic acid (HA) biosynthetic operon was chromosomally inserted with an efficiency of 69%. Furthermore, repression of a heterologous reporter gene was achieved, demonstrating the versatility of the tool kit. The performance of our tool kit is comparable with those of systems developed for Escherichia coli and Saccharomyces cerevisiae, which rely on replicating vectors to implement CRISPR-Cas9 machinery. IMPORTANCE: In this paper, as the first approach, we report implementation of the CRISPR-Cas9 system in Bacillus subtilis, which is recognized as a valuable host system for biomanufacturing. The study enables comprehensive engineering of B. subtilis strains with virtually any desired genotypes/phenotypes and biochemical properties for extensive industrial application.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.