Recent progress in molecular diagnostics: The synergy of rolling circle amplification and CRISPR/Cas systems (2018–2024) – A concise review

The fusion of Rolling Circle Amplification (RCA) and the Clustered Regularly Interspaced Short Palindromic Repeats–CRISPR-associated proteins (CRISPR-Cas) stands at the cutting edge of molecular diagnostics, offering a potent tool for the targeted investigation of genetic material. RCA, known for its robust isothermal amplification capabilities, enables the exponential replication of circular nucleic acid templates. The integration of RCA with CRISPR-Cas technologies has paved the way for innovative diagnostic strategies that boast remarkable specificity and sensitivity in the detection of nucleic acid sequences. This review outlines the fundamental aspects of RCA, including its underlying principles, the diversity of its methodologies, and its variants. We provide a comprehensive overview of the latest advancements in the design, development, and practical application of RCA biosensors that employ the CRISPR/Cas system for enhanced biosensing capabilities. The review further explores the broad potential of RCA in applications beyond traditional biological targets, such as protein and biomarker detection. We conclude with an analytical discussion on the present challenges and outlook of RCA-CRISPR/Cas-based diagnostics, underscoring the transformative impact of these technologies on the various fields of molecular diagnostics.