Microchannel-based portable sensor for p53 gene based on CRISPR/Cas12a using multimeter as readout

Abstract

The DNA intensity modified on the surface of the inner wall of microchannel can regulate the resistance of the microchannel and which can be used to design biosensors for diverse targets. In this study, a portable biosensor based on the regulation of the charge density in microchannel using multimeter as redout had been designed for disease related DNA sequence detection (p53 gene had been chosen as model target). The inner wall of microchannel was functionalized with ssDNA firstly, where target p53 activated Cas12a via crRNA, whose cleavage activity altered the inner surface charge density, thereby changing resistance of the channel, which can be detected by a simple multimeter easily. Key operational parameters such as KCl concentration, CRISPR/Cas12a cleavage time, the concentration of modified ssDNA, and the amounts of Cas12a/crRNA complex were systematically optimized to enhance detection performance. Under optimized conditions, the changing of the resistance of the channel exhibited a linear relationship with the logarithm of p53 gene concentration ranging from 50 fM to 50 pM, achieving a detection limit of 15 fM (S/N = 3). The sensor demonstrated high sensitivity, specificity, and operational convenience, offering a promising point-of-care testing for disease related DNA detection.