CRISPR/Cas13a-triggered entropy-driven amplification for colorimetric and fluorescent dual-mode detection of microRNA.
Anal Biochem
; 689: 115499, 2024 Jun.
Article
em En
| MEDLINE
| ID: mdl-38431141
ABSTRACT
MicroRNAs (miRNAs) are crucial biomarkers for the early detection and monitoring of disease progression of chronic obstructive pulmonary disease (COPD). Herein, we have devised a method for detecting miRNA using a combination of colorimetric and graphene oxide-based fluorescent techniques. The target miRNA in our design could precisely activate the trans-cleavage activity of the CRISPR-Cas13a system. The activated Cas13a enzyme cuts the "rUrU" section in the P1 probe, generating a nicking site to induce entropy-driven amplification (EDA). One of the available EDA products has the capability to unfold the hairpin probe, thereby initiating the catalytic hairpin assembly, exposing the G-quadruplex structure, facilitating the subsequent color response. The fuel strand labeled with Cy3 successfully established a double-stranded DNA structure with DNA3, and consequently the Cy3 would not be quenched by graphene oxide (GO). The implementation of the dual-mode technique in this method yields greater benefits in terms of improving the precision and consistency of the miRNA measurements. The developed method has the capability to fluorescently measure miRNA-21 levels down to a concentration of 5.8 fM. In addition, the analysis of miRNA targets from clinical samples using this method demonstrates its promising utility in the fields of biomedical research of COPD.
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Texto completo:
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Base de dados:
MEDLINE
Assunto principal:
Técnicas Biossensoriais
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Doença Pulmonar Obstrutiva Crônica
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MicroRNAs
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Grafite
Limite:
Humans
Idioma:
En
Ano de publicação:
2024
Tipo de documento:
Article