A CRISPR-Cas12a-based electrochemical biosensor for the detection of microphthalmia-associated transcription factor

被引:15
作者
Luo, Qisheng [1 ]
Zhang, Chunyuan [1 ]
Deng, Xiandong [1 ]
Liu, Dongyuan [1 ]
Pan, Xingchen [1 ]
Gong, Yuanxun [1 ]
Tang, Qianli [1 ]
Zhang, Kai [2 ]
Liao, Xianjiu [3 ]
机构
[1] Youjiang Med Univ Nationalities, Affiliated Hosp, Baise 533000, Guangxi, Peoples R China
[2] Nanjing Univ Informat Sci & Technol, Sch Chem & Mat Sci, Nanjing 210044, Peoples R China
[3] West Guangxi Key Lab Prevent & Treatment High Inci, Baise 533000, Guangxi, Peoples R China
基金
中国国家自然科学基金;
关键词
MITF; CRISPR-Cas12a system; Hairpin DNA; Electrochemical biosensor; Differential pulse voltammetry; SENSITIVE DETECTION; PROBE; ASSAY;
D O I
10.1007/s00604-023-06164-5
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
A novel electrochemical biosensor that combines the CRISPR-Cas12a system with a gold electrode is reported for the rapid and sensitive detection of microphthalmia-associated transcription factor (MITF). The biosensor consists of a gold electrode modified with DNA1, which contains the target sequence of MITF and is labeled with ferrocene, an electroactive molecule. The biosensor also includes hairpin DNA, which has a binding site for MITF and can hybridize with helper DNA to form a double-stranded complex that activates CRISPR-Cas12a. When MITF is present, it binds to hairpin DNA and prevents its hybridization with helper DNA, thus inhibiting CRISPR-Cas12a activity and preserving the DPV signal of ferrocene. When MITF is absent, hairpin DNA hybridizes with helper DNA and activates CRISPR-Cas12a, which cleaves DNA1 and releases ferrocene, thus reducing the DPV signal. The biosensor can detect MITF with high sensitivity (with an LOD of 8.14 fM), specificity, and accuracy in various samples, such as cell nuclear extracts and human serum. The biosensor can also diagnose and monitor melanocyte-related diseases and melanin production. This work provides a simple, fast, sensitive, and cost-effective biosensor for MITF detection and a valuable tool for applications in genetic testing, disease diagnosis, and drug screening.
引用
收藏
页数:10
相关论文
共 36 条
[1]   A general approach to the design of allosteric, transcription factor-regulated DNAzymes [J].
Adornetto, G. ;
Porchetta, A. ;
Palleschi, G. ;
Plaxco, K. W. ;
Ricci, F. .
CHEMICAL SCIENCE, 2015, 6 (07) :3692-3696
[2]   Real-Time Detection of Transcription Factors Using Target-Converted Helicase-Dependent Amplification Assay with Zero-Background Signal [J].
Cao, Anping ;
Zhang, Chun-yang .
ANALYTICAL CHEMISTRY, 2013, 85 (04) :2543-2547
[3]   Rolling circle amplification based on signal-enhanced electrochemical DNA sensor for ultrasensitive transcription factor detection [J].
Deng, Keqin ;
Li, Chunxiang ;
Huang, Haowen ;
Li, Xiaofang .
SENSORS AND ACTUATORS B-CHEMICAL, 2017, 238 :1302-1308
[4]   Nanoparticle-based fluorescence probe for detection of NF-κB transcription factor in single cell via steric hindrance [J].
Ding, Yuedi ;
Fan, Zhenqiang ;
Yao, Bo ;
Xu, Dong ;
Xie, Minhao ;
Zhang, Kai .
MICROCHIMICA ACTA, 2021, 188 (07)
[5]   Electrochemiluminescence platform for transcription factor diagnosis by using CRISPR-Cas12a trans-cleavage activity [J].
Fan, Zhenqiang ;
Ding, Yuedi ;
Yao, Bo ;
Wang, Jiaying ;
Zhang, Kai .
CHEMICAL COMMUNICATIONS, 2021, 57 (65) :8015-8018
[6]   Microphthalmia-associated transcription factor (MITF) is required but is not sufficient to induce the expression of melanogenic genes [J].
Gaggioli, C ;
Buscà, R ;
Abbe, P ;
Ortonne, JP ;
Ballotti, R .
PIGMENT CELL RESEARCH, 2003, 16 (04) :374-382
[7]   Microphthalmia transcription factor as a molecular marker for circulating tumor cell detection in blood of melanoma patients [J].
Koyanagi, K ;
O'Day, SJ ;
Gonzalez, R ;
Lewis, K ;
Robinson, WA ;
Amatruda, TT ;
Kuo, C ;
Wang, HJ ;
Milford, R ;
Morton, DL ;
Hoon, DSB .
CLINICAL CANCER RESEARCH, 2006, 12 (04) :1137-1143
[8]   Single-cell multiomics: technologies and data analysis methods [J].
Lee, Jeongwoo ;
Hyeon, Do Young ;
Hwang, Daehee .
EXPERIMENTAL AND MOLECULAR MEDICINE, 2020, 52 (09) :1428-1442
[9]   Detecting transcription factors with allosteric DNA-Silver nanocluster switches [J].
Li, Bingzhi ;
Chen, Yue ;
Wang, Jing ;
Lu, Qiaoyun ;
Zhu, Wanying ;
Luo, Jieping ;
Hong, Junli ;
Zhou, Xuemin .
ANALYTICA CHIMICA ACTA, 2019, 1048 :168-177
[10]   A dumbell probe-mediated rolling circle amplification strategy for highly sensitive transcription factor detection [J].
Li, Chunxiang ;
Qiu, Xiyang ;
Hou, Zhaohui ;
Deng, Keqin .
BIOSENSORS & BIOELECTRONICS, 2015, 64 :505-510