Affiliation:
1. Department of Biomedical Engineering City University of Hong Kong Hong Kong China
2. Hong Kong Centre for Cerebro‐cardiovascular Health Engineering Hong Kong Science Park Hong Kong China
3. City University of Hong Kong Shenzhen Research Institute Shenzhen China
Abstract
AbstractDNA glycosylases are a group of enzymes that play a crucial role in the DNA repair process by recognizing and removing damaged or incorrect bases from DNA molecules, which maintains the integrity of the genetic information. The abnormal expression of uracil‐DNA glycosylase (UDG), one of significant DNA glycosylases in the base‐excision repair pathway, is linked to numerous diseases. Here, we proposed a simple UDG activity detection method based on toehold region triggered CRISPR/Cas12a trans‐cleavage. The toehold region on hairpin DNA probe (HP) produced by UDG could induce the trans‐cleavage of ssDNA with fluorophore and quencher, generating an obvious fluorescence signal. This protospacer adjacent motif (PAM)‐free approach achieves remarkable sensitivity and specificity in detecting UDG, with a detection limit as low as 0.000368 U mL−1. Moreover, this method is able to screen inhibitors and measure UDG in complex biological samples. These advantages render it highly promising for applications in clinical diagnosis and drug discovery.
Funder
Science, Technology and Innovation Commission of Shenzhen Municipality
City University of Hong Kong