A universal CRISPR-Cas14a responsive triple-sensitized upconversion photoelectrochemical sensor

Abstract

AbstractIt has recently been discovered that, like other members of the Cas family (12a and 13a), the clustered regularly interspaced short palindrome repeat CRISPR-Cas14a system not only mediates high-sensitivity detection with exceptionally strong gene editing ability but is also generally useful for DNA detection via fluorescence. Photoelectrochemical (PEC) sensors have been widely applied as efficient analytical tools. Measuring electrical signals is more cost-effective and the necessary equipment is more easily portable than fluorescence signal detectors, but their stability still needs to be improved. The high base resolution of CRISPR-Cas14a can compensate for such shortcomings. Therefore, electrical signals and fluorescence signals were combined, and the development of a universal CRISPR-Cas14a-responsive ultrasensitive upconversion PEC sensor is described in this paper. Moreover, strand displacement amplification (SDA) and a near-infrared (NIR) light source were utilized to further improve the stability and sensitivity of the photoelectric signals. At the same time, the modified working electrode (UCNPs-ssDNA-CdS@Au/ITO) on the three-electrode disposable sensor was used as the reporter probe, which cooperates with the trans-cleavage activity of Cas14a endonuclease. To verify the universality of this sensor, the UCNPs-Cas14a-based PEC sensor was applied for the detection of the small-molecule toxin T2 and protein kinase PTK7. Here, we report that the limit of detection of this reagent was within the fg range, successfully applied to the detection of T2 in oats and PTK7 in human serum. We propose that by combining PEC and CRISPR-14a, UCNPs-Cas14a-based PEC sensors could become powerful drivers for the extensive development of ultrasensitive, accurate and cost-effective universal sensors for detection and diagnosis.