CRISPR/Cas13a-assisted amplification-free miRNA biosensor via dark-field imaging and magnetic gold nanoparticles DOI Creative Commons
Jae‐Jun Kim, Jae‐Sang Hong, Hyunho Kim

et al.

Sensors & Diagnostics, Journal Year: 2024, Volume and Issue: 3(8), P. 1310 - 1318

Published: Jan. 1, 2024

MicroRNAs (miRNAs) are short (about 18-24 nucleotides) non-coding RNAs and have emerged as potential biomarkers for various diseases, including cancers. Due to their lengths, the specificity often becomes an issue in conventional amplification-based methods. Next-generation sequencing techniques could be alternative, but long analysis time expensive costs make them less suitable routine clinical diagnosis. Therefore, it is essential develop a rapid, selective, accurate miRNA detection assay using simple, affordable system. In this work, we report CRISPR/Cas13a-based biosensing point-of-care dark-field (DF) imaging. We utilized magnetic-gold nanoparticle (MGNPs) complexes signal probes, which consist of 200 nm-sized magnetic beads 60 gold nanoparticles (AuNPs) linked by DNA hybridization. Once CRISPR/Cas13a system recognized target miRNAs (miR-21-5p), activated Cas13a cleaved bridge linker containing RNA sequences, releasing nm-AuNPs detected quantified portable DF imaging The combination CRISPR/Cas13a, MGNPs, demonstrated amplification-free miR-21-5p within 30 min at limit 500 attomoles (25 pM) with single-base specificity. CRISPR/Cas13a-assisted MGNP-DF achieved simple equipment, thus providing application cancer

Language: Английский

‘Splice-at-will’ Cas12a crRNA engineering enabled direct quantification of ultrashort RNAs DOI Creative Commons

Xinrui Fei,

Chao Lei, Wei Ren

et al.

Nucleic Acids Research, Journal Year: 2025, Volume and Issue: 53(2)

Published: Jan. 11, 2025

Abstract We present a robust ‘splice-at-will’ CRISPR RNA (crRNA) engineering mechanism that overcomes the limitations of clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system in directly detecting ultrashort RNAs. In this strategy, an intact Cas12a crRNA can be split from almost any site spacer region to obtain truncated (tcrRNA) cannot activate even after binding auxiliary DNA activator. While splicing tcrRNAs with moiety RNA, formed combination work together efficiently, enabling engineering. Importantly, exhibits same trans-cleavage activation efficiency as conventional crRNA. Therefore, by rationally designing activator conserved tcrRNA-complementary sequence and arbitrary RNA-of-interest recognition domain, general sensing is established utilizes traditional DNA-activated detect This strategy could faithfully sequences 6–8 nt, which achieved Cas13a systems. Additionally, through flexible design, our method precisely distinguish single-base differences microRNA other sequences. has significantly expanded Cas12a-based diagnostic toolbox opened new avenues for detection.

Language: Английский

Citations

2
Quantum Dot-Based Molecular Beacons for Quantitative Detection of Nucleic Acids with CRISPR/Cas(N) Nucleases DOI
Christopher M. Green, Joseph R. Spangler, Kimihiro Susumu

et al.

ACS Nano, Journal Year: 2022, Volume and Issue: 16(12), P. 20693 - 20704

Published: Nov. 15, 2022

Strategies utilizing the CRISPR/Cas nucleases Cas13 and Cas12 have shown great promise in development of highly sensitive rapid diagnostic assays for detection pathogenic nucleic acids. The most common approaches fluorophore-quencher molecular beacons require strand amplification strategies or optical setups to overcome limitations readout. Here, we demonstrate a flexible strategy assembling luminescent colorimetric quantum dot-nucleic acid hairpin (QD-HP) use diagnostics. This utilizes chimeric peptide-peptide (peptide-PNA) conjugate fluorescently labeled DNA RNA hairpins ZnS-coated QDs. QDs are particularly promising alternatives due their greater brightness, strong UV absorbance with large emission offset, exceptional photostability, potential multiplexing sharp peaks. Using Förster resonance energy transfer (FRET), developed ratiometric reporters capable pM target (without nucleotide amplification) both RNA, further demonstrated capabilities camera-phone detection. flexibility this system is imparted by dual functionality QD as FRET donor central nanoscaffold arranging acids fluorescent acceptors on its surface. method also provides generalized approach that could be applied other nuclease systems.

Language: Английский

Citations

44
Recent progress in nucleic acid detection with CRISPR DOI Open Access

Frank X. Liu,

Johnson Q. Cui, Zhihao Wu

et al.

Lab on a Chip, Journal Year: 2023, Volume and Issue: 23(6), P. 1467 - 1492

Published: Jan. 1, 2023

Critical development of CRISPR-based diagnostics coupled with nucleic acid amplification and amplification-free techniques; various purposes CRISPR including determination, quantification, multiplexed point-of-care diagnostics.

Language: Английский

Citations

22
Non-canonical CRISPR/Cas12a-based technology: A novel horizon for biosensing in nucleic acid detection DOI

Xueying Lei,

Shengnan Cao, Tao Liu

et al.

Talanta, Journal Year: 2024, Volume and Issue: 271, P. 125663 - 125663

Published: Jan. 16, 2024

Language: Английский

Citations

11
MicroRNA Sensors Based on CRISPR/Cas12a Technologies: Evolution From Indirect to Direct Detection DOI
Songcheng Yu,

Xueying Lei,

Chenling Qu

et al.

Critical Reviews in Analytical Chemistry, Journal Year: 2024, Volume and Issue: unknown, P. 1 - 17

Published: March 15, 2024

MicroRNA (miRNA) has emerged as a promising biomarker for disease diagnosis and potential therapeutic targets drug development. The detection of miRNA can serve noninvasive tool in diseases predicting prognosis. CRISPR/Cas12a system great nucleic acid due to its high sensitivity specificity, which been developed be versatile acid-based various fields. However, conversion from RNA DNA with or without amplification operation is necessary based on system, because dsDNA containing PAM sequence ssDNA traditionally considered the activator Cas12a. Until recently, direct by reported. In this review, we provide an overview evolution biosensors indirect direct, would beneficial development CRISPR/Cas12a-based sensors better performance miRNA.

Language: Английский

Citations

8
Comprehensive insights into pathogenicity and emerging CRISPR-integrated biosensors for the early detection of breast cancer biomarkers DOI

Ashok Saini,

Neeraj Dilbaghi, Neelam Yadav

et al.

Microchemical Journal, Journal Year: 2025, Volume and Issue: unknown, P. 112908 - 112908

Published: Jan. 1, 2025

Language: Английский

Citations

1
Photoswitchable CRISPR/Cas12a-Amplified and Co3O4@Au Nanoemitter Based Triple-Amplified Diagnostic Electrochemiluminescence Biosensor for Detection of miRNA-141 DOI
Qian Wang,

Zuhao Zhang,

Lu Zhang

et al.

ACS Applied Materials & Interfaces, Journal Year: 2022, Volume and Issue: 14(29), P. 32960 - 32969

Published: July 15, 2022

In this work, a CRISPR/Cas12a initiated switchable ternary electrochemiluminescence (ECL) biosensor combined with Co3O4@Au nanoemitter is presented for the in vitro monitoring of miRNA-141. Benefiting from advantages high-throughput cargo payload capability and superconductivity, three-dimensional reduced graphene oxide (3D-rGO) was designated as an introductory conducting stratum paper working electrode (PWE). With collaborative participation NPs, transmutation TPrA Ru(bpy)32+/TPrA system can be riotously expedited into exorbitant free radical ions TPrA•, which provoked exaggeration ECL signal. Moreover, programmable enzyme-free hybrid chain reaction (HCR) amplifier on PWE surface accurately anchored assembly nucleic acid tandem accomplished secondary recursion Impressively, multifunctional nonspecific cis/trans-splitting decomposition manipulated photoswitch "on–off" signal state that avoided false-positive diagnosis. The multistrategy cooperative demonstrated extraordinary sensitivity specificity, low detection limit 3.3 fM (S/N = 3) concentration scope 10 to 100 nM, fully corresponded expectation. Overall, innovative methodology paved generous avenue evaluating multifarious biotransformations provided tremendous impetus development real-time diagnosis clinical other biomarkers.

Language: Английский

Citations

35
The CRISPR-Cas system as a tool for diagnosing and treating infectious diseases DOI Open Access

Juan Lou,

Bin Wang, Junwei Li

et al.

Molecular Biology Reports, Journal Year: 2022, Volume and Issue: 49(12), P. 11301 - 11311

Published: July 20, 2022

Language: Английский

Citations

33
High-Throughput μPAD with Cascade Signal Amplification through Dual Enzymes for arsM in Paddy Soil DOI

Haorui Cao,

Kang Mao, Jiajia Yang

et al.

Analytical Chemistry, Journal Year: 2024, Volume and Issue: 96(16), P. 6337 - 6346

Published: April 13, 2024

The

Language: Английский

Citations

7
Programmable readout sensor for microRNA: CRISPR/Cas12a-assisted multi-amplification strategy activated photoelectrochemistry-colorimetry detection DOI
Haoran Shen, Hui Ying Yang, Aori Qileng

et al.

Sensors and Actuators B Chemical, Journal Year: 2022, Volume and Issue: 371, P. 132585 - 132585

Published: Sept. 1, 2022

Language: Английский

Citations

21