ArticleMethods in molecular biology (Clifton, N.J.)2025
Golden Gate Cloning of Synthetic CRISPR RNA Spacer Sequences.
Article in Methods in molecular biology (Clifton, N.J.), 2025. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
What it found
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Who cites it
2 citing papers in PubMed.
- Type I-Fv and engineered type IV-A1 CRISPR-Cas effectors facilitate genome reduction in Escherichia coli.Nucleic acids research · 2025Article
- Real-time imaging of bacterial colony growth dynamics for cells with Type IV-A1 CRISPR-Cas activity.microLife · 2025Article
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Authors and funding
2 authors.
Funding
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Abstract
Prokaryotes use CRISPR-Cas systems to interfere with viruses and other mobile genetic elements. CRISPR arrays comprise repeated DNA elements and spacer sequences that can be engineered for custom target sites. These arrays are transcribed into precursor CRISPR RNAs (pre-crRNAs) that undergo maturation steps to form individual CRISPR RNAs (crRNAs). Each crRNA contains a single spacer that identifies the target cleavage site for a large variety of Cas protein effectors. Precise manipulation of spacer sequences within CRISPR arrays is crucial for advancing the functionality of CRISPR-based technologies. Here, we describe a protocol for the design and creation of a minimal, plasmid-based CRISPR array to enable the expression of specific, synthetic crRNAs. Plasmids contain entry spacer sequences with two type IIS restriction sites and Golden Gate cloning enables the efficient exchange of these spacer sequences. Factors that influence the compatibility of the CRISPR arrays with native or recombinant Cas proteins are discussed.
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Registered trials
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