Abstract

Clustered regularly interspaced short palindromic repeats (CRISPR)-based field-effect transistor (FET) sensors provide a promising platform for highly specific and amplification-free nucleic acid detection. However, their direct application in identifying target sequences within intact genomic DNA remains underexplored. We developed a CRISPR-Cas9 FET sensor for detecting DNA sequences by immobilizing Cas9 onto a graphene-MXene sensing interface via (3-aminopropyl)triethoxysilane (APTES) functionalization. The sensor targeted Stx1, a 440 bp laboratory-amplified fragment associated with Shiga toxin production in pathogenic E. coli. Gel electrophoresis confirmed the precise cleavage of Stx1, while the nontarget sequence Stx2 remained undetected. FET measurements further validated the sensor's specificity, showing a significantly stronger signal response for Stx1 than Stx2. To evaluate its performance in complex genomic samples, we analyzed four genomic DNA samples containing varying Stx1 concentrations. The sensor distinguished differences in target sequence abundance, showing its robustness for real-world genomic detection. This work demonstrates the feasibility of integrating CRISPR-Cas9 with FET technology for direct, label-free, and cost-effective genomic diagnostics, paving the way for highly specific biosensing applications.

Department(s)

Civil, Architectural and Environmental Engineering

Keywords and Phrases

CRISPR-Cas9; FET; genomic DNAs; specific cleavage; target sequence

International Standard Serial Number (ISSN)

2574-0970

Document Type

Article - Journal

Document Version

Citation

File Type

text

Language(s)

English

Rights

© 2025 American Chemical Society, All rights reserved.

Publication Date

22 Aug 2025

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