CRISPR-/Cas12a-Mediated Liposome-Amplified Strategy for the Surface-Enhanced Raman Scattering and Naked-Eye Detection of Nucleic Acid and Application to Food Authenticity Screening

Abstract

Surface-enhanced Raman scattering (SERS) has been recognized as a powerful tool for biosensors due to the ultrahigh sensitivity and unique fingerprint information. However, there are some limitations in trace target nucleic acid detection for the restricted signal-transducing and amplification strategies. Inspired by CRISPR/Cas12a with specific target DNA-activated collateral single-strand DNA (ssDNA) cleavage activity and liposome with signal molecule-loading properties, we first proposed a sensitive SERS-based on-site nucleic acid detection strategy mediated by CRISPR/Cas12a with trans-cleavage activity on ssDNA linkers utilized to capture liposomes. Liposomes loading two kinds of signal molecules, 4-nitrothiophenol (4-NTP) and cysteine, could achieve the dual-mode detection of target DNA with SERS and naked eye, respectively. The promptly amplified signals were initiated by the triggered breakdown of signal molecule-loaded liposomes. Emancipated 4-NTP, a biological-silent Raman reporter, would achieve highly selective and sensitive SERS measurement. Released cysteine induced the aggregation of plasmonic gold nanoparticles, leading to an obvious red to blue colorimetric shift to realize portable naked-eye detection. With this strategy, target nucleic acid concentration was dexterously converted into SERS and visualization signals and could be detected as low as 100 aM and 10 pM, respectively. The approach was also successfully applied to determine meat adulteration, achieving the detection of a low adulteration ratio in the complicated food matrix. We anticipate that this strategy will not only be regarded as a universal platform for the on-site detection of food authenticity but also broaden SERS application for the accurate determination of diverse biomarkers.

CRISPR‐Cas‐based detection for food safety problems: Current status, challenges, and opportunities

“…The liposome amplification strategy was coupled with CRISPR‐Cas12a and applied to differentiate meat adulteration. A low adulteration ratio could be achieved even in the complicated food matrices (Liu, Chen, et al., 2021). The detection signals in this method can be collected by the naked eye without interference from complex foods and the detection is at a low adulteration rate.…”

Innovative Application of SERS in Food Quality and Safety: A Brief Review of Recent Trends

“…As vibrational spectroscopic techniques, Raman/SERS have the potential to fulfill the industrial need for food quality and authenticity analyses [ 56 ]. Two strategies to judge authenticity include: (1) combination with chemometrics to achieve the identification and quantification of food samples; (2) combination with specific target DNA-modified SERS substrates with a target-responsive Raman dye that can recognize the target ( Table 7 ) [ 58 , 59 ]. The SERS biosensors are regarded as a universal platform for the on-site determination of food quality, authenticity, and safety [ 60 ].…”

Nanotechnology in food and water security: on-site detection of agricultural pollutants through surface-enhanced Raman spectroscopy

“…Illegal adulteration is not obtained only by the addition of chemical adulterants but also by gene editing. As an adulteration process, Liu et al [ 75 ] detected not a direct pollutant but an adulterated duck gene using SERS (Fig. 4C ).…”