Detection of Bacterial Genomic DNA Fragments Using the MXene CRISPR-Cas9 Field-Effect Transistor

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.