A FEN 1-driven DNA walker-like reaction coupling with magnetic bead-based separation for specific SNP detection

Shijie Xu; Jian Chen; Fang Yang; Zhihao Yang; Jianrong Xu; Lanyue Wang; Lina Bian; Lihua Liu; Xiaoyu Zhao; Yunshan Zhang

doi:10.3389/fbioe.2023.1279473

A FEN 1-driven DNA walker-like reaction coupling with magnetic bead-based separation for specific SNP detection

Front Bioeng Biotechnol. 2023 Nov 3:11:1279473. doi: 10.3389/fbioe.2023.1279473. eCollection 2023.

Authors

Shijie Xu¹, Jian Chen¹, Fang Yang¹, Zhihao Yang², Jianrong Xu³, Lanyue Wang¹, Lina Bian², Lihua Liu¹, Xiaoyu Zhao^{2

4}, Yunshan Zhang²

Affiliations

¹ School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan, Hunan, China.
² Research Center for Intelligent Sensing Systems, Zhejiang Laboratory, Hangzhou, China.
³ School of Electronic Engineering, Nanjing Normal University, Taizhou College, Taizhou, China.
⁴ College of Materials and Environmental Engineering, Hangzhou Dianzi University, Hangzhou, China.

Abstract

Single-nucleotide polymorphism (SNP) plays a key role in the carcinogenesis of the human genome, and understanding the intrinsic relationship between individual genetic variations and carcinogenesis lies heavily in the establishment of a precise and sensitive SNP detection platform. Given this, a powerful and reliable SNP detection platform is proposed by a flap endonuclease 1 (FEN 1)-driven DNA walker-like reaction coupling with a magnetic bead (MB)-based separation. A carboxyfluorescein (FAM)-labeled downstream probe (DP) was decorated on a streptavidin magnetic bead (SMB). The target DNA, as a walker strand, was captured by hybridization with DP and an upstream probe (UP) to form a three-base overlapping structure and execute the walking function on the surface of SMB. FEN 1 was employed to specifically recognize the three-base overlapping structure and cut the 5'flap at the SNP site to report the walking event and signal amplification. Considering the fact that the fluorescence was labeled on the cleavage and uncleavage sequences of DP and the target DNA-triggered walking event was undistinguishable from the mixtures, magnetic separation came in handy for cleavage probe (CP) isolation and discrimination of the amplified signal from the background signal. In comparison with the conventional DNA walker reaction, this strategy was coupling with SMB-based separation, thus promising a powerful and reliable method for SNP detection and signal amplification.

Keywords: DNA walker-like reaction; KRAS gene; flap endonuclease 1; single-nucleotide polymorphism; streptavidin magnetic beads.

Grants and funding

The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was supported by the Key Research Project (2022MH0AC01), the Natural Science Foundation of Zhejiang Province, China (No. 665LQ23C050004), the Natural Science Foundation of Hunan Province, China (Grant No. 2020JJ5156), and the Fundamental Research Fund for the Provincial Universities of Zhejiang (No. GK239909299001-007).