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

清脆的 核酸 核酸检测 化学 计算生物学 生物 生物化学 基因
作者
Cia‐Hin Lau,Siping Huang,Haibao Zhu
出处
期刊:Critical Reviews in Biotechnology [Informa]
卷期号:: 1-28
标识
DOI:10.1080/07388551.2024.2399560
摘要

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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