Amplification-free nucleic acids detection with next-generation CRISPR/dx systems.

IF 8.1 2区 工程技术 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Critical Reviews in Biotechnology Pub Date : 2024-09-22 DOI:10.1080/07388551.2024.2399560
Cia-Hin Lau, Siping Huang, Haibao Zhu
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Abstract

CRISPR-based diagnostics (CRISPR/Dx) have revolutionized the field of molecular diagnostics. It enables home self-test, field-deployable, and point-of-care testing (POCT). Despite the great potential of CRISPR/Dx in diagnoses of biologically complex diseases, preamplification of the template often is required for the sensitive detection of low-abundance nucleic acids. Various amplification-free CRISPR/Dx systems were recently developed to enhance signal detection at sufficient sensitivity. Broadly, these amplification-free CRISPR/Dx systems are classified into five groups depending on the signal enhancement strategies employed: CRISPR/Cas12a and/or CRISPR/Cas13a are integrated with: (1) other catalytic enzymes (Cas14a, Csm6, Argonaute, duplex-specific nuclease, nanozyme, or T7 exonuclease), (2) rational-designed oligonucleotides (multivalent aptamer, tetrahedral DNA framework, RNA G-quadruplexes, DNA roller machine, switchable-caged guide RNA, hybrid locked RNA/DNA probe, hybridized cascade probe, or "U" rich stem-loop RNA), (3) nanomaterials (nanophotonic structure, gold nanoparticle, micromotor, or microbeads), (4) electrochemical and piezoelectric plate biosensors (SERS nanoprobes, graphene field-effect transistor, redox probe, or primer exchange reaction), or (5) cutting-edge detection technology platforms (digital bioanalysis, droplet microfluidic, smartphone camera, or single nanoparticle counting). Herein, we critically discuss the advances, pitfalls and future perspectives for these amplification-free CRISPR/Dx systems in nucleic acids detection. The continued refinement of these CRISPR/Dx systems will pave the road for rapid, cost-effective, ultrasensitive, and ultraspecific on-site detection without resorting to target amplification, with the ultimate goal of establishing CRISPR/Dx as the paragon of diagnostics.

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利用新一代 CRISPR/dx 系统进行无扩增核酸检测。
基于 CRISPR 的诊断(CRISPR/Dx)彻底改变了分子诊断领域。它实现了家庭自测、现场部署和护理点检测(POCT)。尽管 CRISPR/Dx 在诊断生物复杂疾病方面具有巨大潜力,但要灵敏地检测低丰度核酸,往往需要对模板进行预扩增。最近开发了多种无扩增 CRISPR/Dx 系统,以提高信号检测的灵敏度。根据所采用的信号增强策略,这些无扩增 CRISPR/Dx 系统大致可分为五类:CRISPR/Cas12a和/或CRISPR/Cas13a与以下方面结合在一起:(1) 其他催化酶(Cas14a、Csm6、Argonaute、双链特异性核酸酶、纳米酶或 T7 外切酶),(2) 合理设计的寡核苷酸(多价aptamer、四面体 DNA 框架、RNA G-四重链、DNA 滚轮机、可切换笼状引导 RNA、杂交锁定 RNA/DNA 探针、杂交级联探针或富含 "U "的茎环 RNA)、(3) 纳米材料(纳米光子结构、金纳米粒子、微马达或微珠),(4) 电化学和压电板生物传感器(SERS 纳米探针、石墨烯场效应晶体管、氧化还原探针或引物交换反应),或 (5) 尖端检测技术平台(数字生物分析、液滴微流控、智能手机摄像头或单纳米粒子计数)。在此,我们将认真讨论这些无扩增 CRISPR/Dx 系统在核酸检测方面的进展、缺陷和未来前景。这些CRISPR/Dx系统的不断完善将为实现快速、经济、超灵敏和超特异的现场检测铺平道路,而无需借助目标扩增,最终目标是将CRISPR/Dx打造成诊断领域的典范。
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来源期刊
Critical Reviews in Biotechnology
Critical Reviews in Biotechnology 工程技术-生物工程与应用微生物
CiteScore
20.80
自引率
1.10%
发文量
71
审稿时长
4.8 months
期刊介绍: Biotechnological techniques, from fermentation to genetic manipulation, have become increasingly relevant to the food and beverage, fuel production, chemical and pharmaceutical, and waste management industries. Consequently, academic as well as industrial institutions need to keep abreast of the concepts, data, and methodologies evolved by continuing research. This journal provides a forum of critical evaluation of recent and current publications and, periodically, for state-of-the-art reports from various geographic areas around the world. Contributing authors are recognized experts in their fields, and each article is reviewed by an objective expert to ensure accuracy and objectivity of the presentation.
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