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ORIGINAL RESEARCH article

Front. Bioeng. Biotechnol.

Sec. Nanobiotechnology

Volume 13 - 2025 | doi: 10.3389/fbioe.2025.1602494

Coupling Enzymatic Digestion with Nanopore Sensing for Low-Abundance EGFR-L858R Mutation Detection

Provisionally accepted
Ling  YanLing Yan1*Zhenxin  WangZhenxin Wang2Yajie  YinYajie Yin2Changchun  NiuChangchun Niu1Yang  LuoYang Luo1
  • 1Chongqing Medical University, Chongqing, China
  • 2Chongqing Institute of Green and Intelligent Technology, Chinese Academy of Sciences (CAS), Chongqing, Chongqing Municipality, China

The final, formatted version of the article will be published soon.

Single nucleotide variants (SNVs) serve as crucial biomarkers for tumor diagnosis, treatment, and prognosis. However, their detection remains challenging due to the low mutation frequency and limited abundance. In this study, we developed a sensitive and efficient SNV detection method combining T7 endonuclease I (T7E1) digestion and nanopore detection. The assay was designed to detect the L858R mutation in exon 21 of the EGFR gene, a key driver mutation in various cancers. Magnetic bead probes were utilized to capture and purify target DNA, followed by enzymatic cleavage and nanopore detection. The optimal nanopore detection voltage was determined to be 500 mV, ensuring a clear and reproducible signal for identifying digested DNA fragments. This approach enabled the detection of L858R mutations with a sensitivity as low as 0.1%. Compared to conventional sequencing and digital PCR, our method is simple, time-efficient, and cost-effective. This strategy offers a promising platform for SNV detection, providing a valuable tool for tumor diagnosis and personalized treatment.

Keywords: Single nucleotide variant, EGFR L858R, T7E1 digestion, Nanopore detection, Magnetic bead separation, tumor biomarker

Received: 29 Mar 2025; Accepted: 21 Aug 2025.

Copyright: © 2025 Yan, Wang, Yin, Niu and Luo. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

* Correspondence: Ling Yan, Chongqing Medical University, Chongqing, China

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