CRISPR/Cas-Powered Amplification-Free Detection of Nucleic Acids: Current State of the Art, Challenges, and...

CRISPR/Cas system is becoming an increasingly influential technology that has been repositioned in nucleic acid detection. A preamplification step is usually required to improve the sensitivity of CRISPR/Cas-based detection. The striking biological features of CRISPR/Cas, including programmability, high sensitivity and sequence specificity, and single-base resolution. More strikingly, the target-activated trans-cleavage could act as a biocatalytic signal transductor and amplifier, thereby empowering it to potentially perform nucleic acid detection without a preamplification step. The reports of such work are on the rise, which is not only scientifically significant but also promising for futuristic end-user applications. This review started with the introduction of the detection methods of nucleic acids and the CRISPR/Cas-based diagnostics (CRISPR-Dx). Next, we objectively discussed the pros and cons of preamplification steps for CRISPR-Dx. We then illustrated and highlighted the recently developed strategies for CRISPR/Cas-powered amplification-free detection that can be realized through the uses of ultralocalized reactors, cascade reactions, ultrasensitive detection systems, or others. Lastly, the challenges and futuristic perspectives were proposed. It can be expected that this work not only makes the researchers better understand the current strategies for this emerging field, but also provides insight for designing novel CRISPR-Dx without a preamplification step to win practicable use in the near future.