Artificial nucleic acid circuits with precisely controllable dynamic and function have shown great promise in biosensing, but their utility in molecular diagnostics is still restrained by the inability to process genomic DNA directly and moderate sensitivity. To address this limitation, we present a CRISPR-Cas–powered catalytic nucleic acid circuit, namely, CRISPR-Cas–only amplification network (CONAN), for isothermally amplified detection of genomic DNA. By integrating the stringent target recognition, helicase activity, and trans-cleavage activity of Cas12a, a Cas12a autocatalysis-driven artificial reaction network is programmed to construct a positive feedback circuit with exponential dynamic in CONAN. Consequently, CONAN achieves one-enzyme, one-step, real-time detection of genomic DNA with attomolar sensitivity. Moreover, CONAN increases the intrinsic single-base specificity of Cas12a, and enables the effective detection of hepatitis B virus infection and human bladder cancer–associated single-nucleotide mutation in clinical samples, highlighting its potential as a powerful tool for disease diagnostics.

A generic sensing strategy that integrates CRISPR-Cas12a with a DNA circuit is proposed for amplified detection of microRNA.

A Ti3C2QD-based fluorescent probe for an ALP activity assay and embryonic stem cell identification was developed by taking advantage of the inner filter effect.

Precise and dynamic imaging of extracellular pH is one crucial yet challenging task for studying cell physiological and pathological processes. Here, we construct a DNA tweezer to dynamically monitor pH changes of cellular microenvironments. The DNA tweezer contains three key elements: a three-strand ssDNA-frame labeled with cholesterol to anchor it on the cell membrane, a pH-sensitive i-motif sequence in the middle to dynamically control the switch between the "open" and "closed" states of the DNA tweezer, and a pair of FRET fluorophores (rhodamine green and rhodamine red) on the two arms of the tweezer to reflect its state. With cholesterol, a natural component of cell membranes, as an anchoring element, the sensor exhibited high cell-membrane-insertion efficiency and low cytotoxicity. Using the i-motif as a sensing element, it can quickly and reversibly respond to extracellular pH in the pH range of 5.0−7.5 and further perform real-time imaging of cell-surface-pH changes with excellent spatial and temporal resolution. Moreover, apoplastic-pH change during the alkalization process of plant roots caused by rapid-alkalinization factor (RALF1) was directly detected by the sensor, demonstrating the potential applications of the sensor in cell biology, biomedical research, and plant-tissue engineering.