Cascade DNA Circuits Mediated CRISPR‐Cas12a Fluorescent Aptasensor based on Multifunctional Fe₃O₄@hollow‐TiO₂@MoS₂ Nanochains for Tetracycline Determination

Abstract

Herein, for the first time, the CRISPR‐Cas12a system is combined with aptamer, cascaded dynamic DNA network circuits, and Fe3O4@hollow‐TiO2@MoS2 nanochains (Fe3O4@h‐TiO2@MoS2 NCs) to construct an efficient sensing platform for tetracycline (TC) analysis. In this strategy, specific recognition of the target is transduced and amplified into H1‐H2 duplexes containing the specific sequence of Cas12a‐crRNA through an aptamer recognition module and the dual amplification dynamic DNA network. Subsequently, the obtained activated Cas12a protein non‐specifically cleaves the adjacent reporter gene ssDNA‐FAM to dissociate the FAM molecule from the quencher Fe3O4@h‐TiO2@MoS2 NCs, resulting in the recovery of the fluorescence signal and further signal amplification. Particularly, the synthesized multifunctional Fe3O4@h‐TiO2@MoS2 NCs composites also exhibit superb magnetic separability and photocatalytic degradation ability. Under optimal conditions, the aptasensor displays a distinct linear relationship with the logarithm of TC concentration, and the limit of detection is as low as 0.384 pg mL−1. Furthermore, the results of spiked recovery confirm the viability of the proposed aptasensor for TC quantification in real samples. This study extends the application of the CRISPR‐Cas12a system in the field of analytical sensing and contributes new insights into the exploration of reliable tools for monitoring and treating hazards in food and environment.

CRISPR/Cas System: The Accelerator for the Development of Non-nucleic Acid Target Detection in Food Safety

“…Furthermore, Mahas et al used the allosteric transcription factor (aTF) to regulate the expression of a CRISPR array coupled with Cas12a activity to develop a diagnosis platform for the detection of different tetracycline antibiotics with high sensitivity and specificity, using TetR as the aTF . Cascaded dynamic DNA network circuits and Fe 3 O 4 @hollow-TiO 2 @MoS 2 nanochains (Fe 3 O 4 @hollow-TiO 2 @MoS 2 NCs) were applied to develop an efficient analysis platform for tetracycline (TC) . In the system, the recognized target was transduced and amplified through an aptamer recognition module and a double amplification dynamic DNA network, EDC-CHA (entropy-driven catalysis), forming a trigger for Cas12a.…”

“…82 Cascaded dynamic DNA network circuits and Fe 3 O 4 @hollow-TiO 2 @ MoS 2 nanochains (Fe 3 O 4 @hollow-TiO 2 @MoS 2 NCs) were applied to develop an efficient analysis platform for tetracycline (TC). 83 In the system, the recognized target was transduced and amplified through an aptamer recognition module and a double amplification dynamic DNA network, EDC-CHA (entropy-driven catalysis), forming a trigger for Cas12a. Cas12 was then activated to nonspecifically cleave the nearby reporter ssDNA-FAM, causing the FAM molecule to dissociate from the quenching agent Fe 3 O 4 @hollow-TiO 2 @MoS 2 NCs, which resulted in recovery of fluorescence signals and further signal enhancement.…”

Paper-Supported Photoelectrochemical Biosensor for Dual-Mode miRNA-106a Assay: Integration of Luminescence-Confined Upconversion-Actuated Fluorescent Resonance Energy Transfer and CRISPR/Cas13a-Powered Cascade DNA Circuits

“…In contrast to the onefold circuit, the DNA cascade circuit highly amplifies the target sensing signal. 60 Additionally, DNA has powerful strengths of sequence programmability, making it anticipated for these series-wound DNA circuits to be integrated with the CRISPR/Cas13a strategy to research a novel biosensor. Compared with the ordinary fluorescently labeled UCNPs, the upconversion luminescence (UCL) coordinated with the fluorescent resonance energy transfer (FRET) selecting UCNPs as the energy donators can favorably eliminate the inveracious outputs and reputably facilitate the assaying productivity in complicated organismal environments.…”

“…In designed cascaded DNA circuit assay networks, the produce of the upriver circuit works as a bridgeable connector to actuate the downriver amplification circuit. In contrast to the onefold circuit, the DNA cascade circuit highly amplifies the target sensing signal . Additionally, DNA has powerful strengths of sequence programmability, making it anticipated for these series-wound DNA circuits to be integrated with the CRISPR/Cas13a strategy to research a novel biosensor.…”

Amplified Assembly of G-Quadruplex-Decorated DNA Network Nanostructure toward AIE Signaling-Based Sensitive Biosensing

“…In the biosensing field, labeling methods generally have high specificity and high sensitivity. Compared to typical antibodies, aptamers can be used for the specific recognition of a large variety of target analytes and also possess excellent stability and terminus-modification properties for constructing a broad array of homogeneous labeled biosensing methods. − Furthermore, their functional nucleic acid characteristics also enable the easy development of powerful nucleic acid amplification techniques based on target biorecognition-induced conformational transformations of relevant nucleic acids. − These not only eliminate the need for the sophisticated preparation of functional nanoprobes for signal amplification but also omit the laborious multistep separation and washing manipulations required in traditional biosensing strategies.…”