Allosteric probe-triggered isothermal amplification to activate CRISPR/Cas12a for sensitive electrochemiluminescence detection of Salmonella

Chunyan Wang; Yutian Zhang; Shanshan Liu; Yashi Yin; Gao-Chao Fan; Yizhong Shen; Heyou Han; Wenjing Wang

doi:10.1016/j.foodchem.2023.136382

Allosteric probe-triggered isothermal amplification to activate CRISPR/Cas12a for sensitive electrochemiluminescence detection of Salmonella

Food Chem. 2023 Nov 1:425:136382. doi: 10.1016/j.foodchem.2023.136382. Epub 2023 May 24.

Authors

Chunyan Wang¹, Yutian Zhang¹, Shanshan Liu¹, Yashi Yin¹, Gao-Chao Fan², Yizhong Shen³, Heyou Han⁴, Wenjing Wang⁵

Affiliations

¹ State Key Laboratory of Agricultural Microbiology, College of Science, Huazhong Agricultural University, Wuhan 430070, China.
² Shandong Key Laboratory of Biochemical Analysis, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, China.
³ School of Food & Biological Engineering, Key Laboratory for Agricultural Products Processing of Anhui Province, Hefei University of Technology, Hefei 230009, China. Electronic address: yzshen@hfut.edu.cn.
⁴ State Key Laboratory of Agricultural Microbiology, College of Science, Huazhong Agricultural University, Wuhan 430070, China. Electronic address: hyhan@mail.hzau.edu.cn.
⁵ State Key Laboratory of Agricultural Microbiology, College of Science, Huazhong Agricultural University, Wuhan 430070, China. Electronic address: wjwang@mail.hzau.edu.cn.

PMID: 37276664
DOI: 10.1016/j.foodchem.2023.136382

Abstract

We report an electrochemiluminescence (ECL) sensor for Salmonella detection based on allosteric probe as a bio-recognition element and CRISPR/Cas12a as a signal amplification strategy. In the presence of Salmonella, the structure switching occurs on allosteric probes, resulting in their hybridization with primers to trigger isothermal amplification. Salmonella is then released to initiate the next reaction cycle accompanying by generating a large amount of dsDNA, which are subsequently recognized by CRISPR-gRNA for activating the trans-cleavage activity of Cas12a. Furthermore, the activated Cas12a can indiscriminately cut the ssDNA which is bound to the electrode, enabling the release of the ECL emitter porphyrinic Zr metal - organic framework (MOF, PCN-224) and exhibiting a decreased ECL signal accordingly. The linear range is 50 CFU·mL^-1-5 × 10⁶ CFU·mL^-1 and the detection limit is calculated to be 37 CFU·mL^-1. This method sensitively detects Salmonella in different types of real samples, indicating it is a promising strategy for Salmonella detection.

Keywords: Aptamer; CRISPR/Cas12a; Electrochemiluminescence; Isothermal amplification; PCN-224; Salmonella.

MeSH terms

Biosensing Techniques*
CRISPR-Cas Systems
DNA Primers
DNA, Single-Stranded
Electrodes
Metal-Organic Frameworks*
Salmonella / genetics

Substances

DNA Primers
DNA, Single-Stranded
Metal-Organic Frameworks