A Label-Free Photoelectrochemical Biosensor Based on CRISPR/Cas12a System Responsive Deoxyribonucleic Acid Hydrogel and “Click” Chemistry

Abstract

A novel label-free photoelectrochemical (PEC) biosensor is presented in this work. As a barrier, the DNA hydrogel could block the coupling between g-C 3 N 4 and CdS quantum dots (QDs). Therefore, extremely low photocurrent signals were obtained. The presence of target microRNA-21 can initiate the rolling circle amplification (RCA) reaction, which in turn produces many repeated sequences to activate the CRISPR/Cas12a system. The trans-cleavage activity of the CRISPR/Cas12a system led to the degradation of DNA hydrogels efficiently. As a result, the g-C 3 N 4 /CdS QDs heterojunction was formed through "click" chemistry. Through the amplification of the RCA and CRISPR/Cas12a system, the sensitivity of the PEC biosensor was improved significantly with the detection limit of 3.2 aM. The proposed sensor also showed excellent selectivity and could be used to detect actual samples. In addition, the modular design could facilitate the detection of different objects. Thus, the proposed CRISPR/ Cas12a system responsive DNA hydrogel provides a simple, sensitive, and flexible way for label-free PEC analysis.

Chemiluminescence Sensor for miRNA-21 Detection Based on CRISPR-Cas12a and Cation Exchange Reaction

“…Dai et al constructed a universal electrochemical biosensor based on CRISPR-Cas12a system . Our group developed a photoelectrochemical sensor using Cas12a-responsive DNA hydrogel . However, there have been few reports on the use of CRISPR technology for CL sensors. , …”

“…34 Our group developed a photoelectrochemical sensor using Cas12a-responsive DNA hydro- gel. 35 However, there have been few reports on the use of CRISPR technology for CL sensors. 36,37 Here, we used the CRISPR-Cas12a system in cation exchange reaction-based CL amplification (CRISPR-CXCLAmp).…”

CRISPR/Cas systems combined with DNA nanostructures for biomedical applications

“…55 The construction principle of DNA hydrogel involved in this strategy is similar to the above work. [49][50][51] The difference is that the DNA hydrogel is encapsulated with platinum nanoparticle decorated metal organic framework nanosheets (PtNPs/Cu TCP (Fe)). The activation of the CRISPR/Cas14a system is achieved through exponential amplification (EXPAR) of activator DNA (cDNA).…”

Ultrasensitive Detection of miRNA via CRISPR/Cas12a Coupled with Strand Displacement Amplification Reaction

“…Recently, clustered regularly interspaced short palindromic repeats (CRISPR) and the CRISPR-associated (Cas) proteins, together referred to as the CRISPR/Cas system, have become an incredibly versatile tool for target recognition and amplification due to their rapid response, excellent recognition specificity, mild reaction temperature, and ideal programmable ability. − Among the Cas proteins family, Cas12a exhibits high enzymatic turnover rate and signal amplification ability, which is able to specifically recognize the target double-stranded DNA (dsDNA) or single-stranded DNA (ssDNA) under the guidance of CRISPR RNA (crRNA) and then trigger both the cleavage of the target DNA ( cis -cleavage) and collateral cleavage of the nontarget ssDNA ( trans -cleavage) . Owing to its characteristic of highly efficient collateral cleavage, the CRISPR/Cas12a system is often used as a tool for signal amplification in biosensors.…”