Modified Unit-Mediated Strand Displacement Reactions for Direct Detection of Single Nucleotide Variants in Active Double-Stranded DNA

作者全名：Yu, Hongyan; Han, Xiaole; Wang, Weitao; Zhang, Yangli; Xiang, Linguo; Bai, Dan; Zhang, Li; Weng, Zhi; Lv, Ke; Song, Lin; Luo, Wang; Yin, Na; Zhang, Yaoyi; Feng, Tong; Wang, Li; Xie, Guoming

作者地址：[Yu, Hongyan; Han, Xiaole; Wang, Weitao; Bai, Dan; Zhang, Li; Lv, Ke; Song, Lin; Luo, Wang; Yin, Na; Zhang, Yaoyi; Feng, Tong; Xie, Guoming] Chongqing Med Univ, Coll Lab Med, Key Lab Clin Lab Diagnost, Chinese Minist Educ, Chongqing 400016, Peoples R China; [Weng, Zhi] Shanghai Jiao Tong Univ, Sch Biomed Engn, State Key Lab Oncogenes & Related Genes, Shanghai 200030, Peoples R China; [Zhang, Yangli; Xiang, Linguo; Wang, Li] Chongqing Med Univ, Affiliated Hosp 1, Ctr Clin Mol Med Detect, Biobank Ctr, Chongqing 400016, Peoples R China

通信作者：Xie, GM (通讯作者)，Chongqing Med Univ, Coll Lab Med, Key Lab Clin Lab Diagnost, Chinese Minist Educ, Chongqing 400016, Peoples R China.; Wang, L (通讯作者)，Chongqing Med Univ, Affiliated Hosp 1, Ctr Clin Mol Med Detect, Biobank Ctr, Chongqing 400016, Peoples R China.

来源：ACS NANO

ESI学科分类：MATERIALS SCIENCE

WOS号：WOS:001226108700001

JCR分区：Q1

影响因子：15.8

年份：2024

卷号：18

期号：19

开始页：12401

结束页：12411

文献类型：Article

关键词：molecular diagnosis; strand-displacement reactions; single nucleotide variations; active double-strandedDNA; PCR

摘要：Accurate identification of single nucleotide variants (SNVs) in key driver genes holds a significant value for disease diagnosis and treatment. Fluorescent probes exhibit tremendous potential in specific, high-resolution, and rapid detection of SNVs. However, additional steps are required in most post-PCR assays to convert double-stranded DNA (dsDNA) products into single-stranded DNA (ssDNA), enabling them to possess hybridization activity to trigger subsequent reactions. This process not only prolongs the complexity of the experiment but also introduces the risk of losing target information. In this study, we proposed two strategies for enriching active double-stranded DNA, involving PCR based on obstructive groups and cleavable units. Building upon this, we explored the impact of modified units on the strand displacement reaction (SDR) and assessed their discriminatory efficacy for mutations. The results showed that detection of low variant allele frequencies (VAF) as low as 0.1% can be achieved. The proposed strategy allowed orthogonal identification of 45 clinical colorectal cancer tissue samples with 100% specificity, and the results were generally consistent with sequencing results. Compared to existing methods for enriching active targets, our approach offers a more diverse set of enrichment strategies, characterized by the advantage of being simple and fast and preserving original information to the maximum extent. The objective of this study is to offer an effective solution for the swift and facile acquisition of active double-stranded DNA. We anticipate that our work will facilitate the practical applications of SDR based on dsDNA.

基金机构：Chongqing Postdoctoral Science Foundation [2022YFC2603800]; National Key R&D Program of China [82172369]; National Natural Science Foundation of China [BJRC202315]; Outstanding Project of Chongqing Medical University [CSTB2022NSCQ-BHX0671]; Chongqing Postdoctoral Science Foundation

基金资助正文：This work was supported by the National Key R&D Program of China (2022YFC2603800), the National Natural Science Foundation of China (82172369), the Outstanding Project of Chongqing Medical University (BJRC202315), and Chongqing Postdoctoral Science Foundation (CSTB2022NSCQ-BHX0671).