Advanced science (Weinheim, Baden-Wurttemberg, Germany)Mar 30, 2026

Using Shape-Sensitive Cas12a with Circular DNA-RNA Guides for Gene Detection Without Amplification and Controlled Gene Editing

CRISPR Gene Editing Weekly Brief ↗PubMed ↗DOI ↗OA ↗

Updated Mar 31, 2026

Abstract

TopCas enables preamplification-free nucleic acid detection and conditional gene editing.

TopCas utilizes circular crRNA to regulate Cas12a's activity, activating it only in the presence of specific target nucleic acids.
The system converts circular crRNA into linear form through a target-activated process, leading to signal amplification.
High specificity and sensitivity in nucleic acid detection are achieved with TopCas, particularly in clinical viral samples.
Conditional gene editing is facilitated by the same mechanism that governs nucleic acid detection.
TopCas shows potential for in vivo real-time molecular imaging, highlighting its versatile applications.

Simplified

Controlled activation of CRISPR-Cas12a is critical for achieving conditional gene editing and molecular diagnostics. As an indispensable component for forming an active complex, CRISPR RNA (crRNA) represents a key route to regulate Cas12a activity. Here, we establish TopCas (Topology-gated Cas12a via DNA-RNA Chimeric Circular crRNA) as a platform for preamplification-free nucleic acid detection and conditional gene editing. Within TopCas, the circular crRNA sterically constrains Cas12a's nuclease activity until target-activated complexes trans-cleave the DNA segment of the chimeric crRNA, converting the circular guide into its linear form and initiating an autocatalytic cascade that culminates in fluorophore release and signal amplification. By the same mechanism, the system conditionally activates Cas12a's gene-editing function (cis-cleavage) exclusively in the presence of specific nucleic acid targets (e.g., viral DNA or RNA). We demonstrate that TopCas affords high specificity and sensitivity in nucleic acid detection, supports accurate detection in clinical viral nucleic acid samples, and shows potential for in vivo real-time molecular imaging, while also demonstrating the feasibility of conditional gene editing. This innovative chimeric circular crRNA-Cas12a system not only provides a new tool for precise disease diagnostics but also offers a promising strategy for personalized therapeutic intervention.

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Funding

Competing interests

The authors declare no conflict of interest.

PubMed

Using Shape-Sensitive Cas12a with Circular DNA-RNA Guides for Gene Detection Without Amplification and Controlled Gene Editing

Abstract

Full Text

Funding

Competing interests

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