Gold Nanomaterials‐Implemented CRISPR‐Cas Systems for Biosensing

Abstract

prokaryotes, [1] which acts as an immune weapon in bacteria to eliminate virus invading. CRISPR-Cas systems developed by bacteria can effectively remove virus genes integrated into bacterial genes. [2,3] The working principle is that under the guidance of RNA, the CRISPR-Cas protein can target the nucleic acid sequence and eliminate it. [4] Inspired by CRISPR-Cas systems, researchers have adapted them to develop gene-editing techniques that have quickly become the most popular technology in life science. [5,6] In addition to the outstanding gene-editing capabilities, CRISPR-Cas systems show great potential as the next-generation techniques for rapid and sensitive biosensing. CRISPR-Cas systems can be easily integrated with a range of nucleic acid-based signal amplification approaches and signal probes because of the precise targeting and cleavage functions toward nucleic acid sequences. Owing to their superior properties including high specificity for identifying single-nucleotide variations, ultrahigh sensitivity, and ease of fabrication, CRISPR-Cas systems can serve as a facile and efficient tool to develop biosensors for point-of-care detections. At present, CRISPR-Cas systems are grouped into class 1 and class 2 systems that are further subdivided into different types and subtypes. Class 1 CRISPR-Cas systems (type I, III, and IV systems) use a wide assortment of smaller Cas proteins (Cas1, Cas2, Cas3, and so on) to form a multisubunit interference complex. On the contrary, class 2 systems (type II, V, and VI systems) use a single, comparatively larger Cas effector protein (Cas9, Cas12a, Cas13a, and so on) for interference and, in certain cases, CRISPR RNA (crRNA) biogenesis. [7] Notably, Cas proteins in class 2 systems have been explored to design CRISPR-Cas-based diagnostic tools, including CRISPR/Cas9-triggered isothermal exponential amplification reaction (CAS-EXPAR), [8] one-hour low-cost multipurpose highly efficient system (HOLMES), [9] HOLMES version 2 (HOLMESv2), [10] specific high-sensitivity enzymatic reporter unlocking (SHERLOCK), [11] SHERLOCK version 2 (SHERLOCKv2), [12] DNA endonuclease-targeted CRISPR transreporter (DETECTR), [13] and so on.Traditional biosensors based on CRISPR-Cas systems generally rely on fluorescent single-stranded DNA (ssDNA) for the readout, while these probes are limited by the low stability and sensitivity in complex samples. To meet these challenges, Due to their superiority in the simple design and precise targeting, clustered regularly interspaced short palindromic repeats (CRISPR)-Cas systems have attracted significant interest for biosensing. On the one hand, CRISPR-Cas systems have the capacity to precisely recognize and cleave specific DNA and RNA sequences. On the other hand, CRISPR-Cas systems such as orthologs of Cas9, Cas12, and Cas13 exhibit cis-cleavage or trans-cleavage activities after recognizing the target sequence. Owing to the cleavage activities, CRISPR-Cas systems can be designed for biosensing by degrading tagged nucleic acids to produce detectable...

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