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- Title
Electrochemical DNA Cleavage Sensing for EcoRV Activity and Inhibition with an ERGO Electrode.
- Authors
Oh, Da Eun; Kim, Hyun Beom; Kim, Tae Hyun
- Abstract
An electrochemically reduced graphene oxide (ERGO) electrode-based electrochemical assay was developed for rapid, sensitive, and straightforward analysis of both activity and inhibition of the endonuclease EcoRV. The procedure uses a DNA substrate designed for EcoRV, featuring a double-stranded DNA (dsDNA) region labeled with methylene blue (MB) and a single-stranded DNA (ssDNA) region immobilized on the ERGO surface. The ERGO electrode, immobilized with the DNA substrate, was subsequently exposed to a sample containing EcoRV. Upon enzymatic hydrolysis, the cleaved dsDNA fragments were detached from the ERGO surface, leading to a decrease in the MB concentration near the electrode. This diminished the electron transfer efficiency for MB reduction, resulting in a decreased reduction current. This assay demonstrates excellent specificity and high sensitivity, with a limit of detection (LOD) of 9.5 × 10−3 U mL−1. Importantly, it can also measure EcoRV activity in the presence of aurintricarboxylic acid, a known inhibitor, highlighting its potential for drug discovery and clinical diagnostic applications.
- Subjects
SINGLE-stranded DNA; DRUG discovery; DNA; EXONUCLEASES; METHYLENE blue; ELECTRODES; ENDONUCLEASES
- Publication
Biosensors (2079-6374), 2024, Vol 14, Issue 2, p73
- ISSN
2079-6374
- Publication type
Article
- DOI
10.3390/bios14020073