An autocatalytic CRISPR-Cas amplification effect propelled by the LNA-modified split activators for DNA sensing.

Publisher: Oxford University Press Country of Publication: England NLM ID: 0411011 Publication Model: Print Cited Medium: Internet ISSN: 1362-4962 (Electronic) Linking ISSN: 03051048 NLM ISO Abbreviation: Nucleic Acids Res Subsets: MEDLINE

Publication: 1992- : Oxford : Oxford University Press
Original Publication: London, Information Retrieval ltd.

CRISPR-Cas Systems* ; Oligonucleotides*/chemistry ; Oligonucleotides*/genetics ; DNA, Single-Stranded*/genetics ; DNA, Single-Stranded*/metabolism ; DNA, Single-Stranded*/chemistry ; Biosensing Techniques*/methods; Humans ; Nucleic Acid Amplification Techniques/methods ; DNA/chemistry ; DNA/genetics ; Cell Line, Tumor ; Catalysis

CRISPR-Cas systems with dual functions offer precise sequence-based recognition and efficient catalytic cleavage of nucleic acids, making them highly promising in biosensing and diagnostic technologies. However, current methods encounter challenges of complexity, low turnover efficiency, and the necessity for sophisticated probe design. To better integrate the dual functions of Cas proteins, we proposed a novel approach called CRISPR-Cas Autocatalysis Amplification driven by LNA-modified Split Activators (CALSA) for the highly efficient detection of single-stranded DNA (ssDNA) and genomic DNA. By introducing split ssDNA activators and the site-directed trans-cleavage mediated by LNA modifications, an autocatalysis-driven positive feedback loop of nucleic acids based on the LbCas12a system was constructed. Consequently, CALSA enabled one-pot and real-time detection of genomic DNA and cell-free DNA (cfDNA) from different tumor cell lines. Notably, CALSA achieved high sensitivity, single-base specificity, and remarkably short reaction times. Due to the high programmability of nucleic acid circuits, these results highlighted the immense potential of CALSA as a powerful tool for cascade signal amplification. Moreover, the sensitivity and specificity further emphasized the value of CALSA in biosensing and diagnostics, opening avenues for future clinical applications.
(© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

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2022YFB3205600 National Key Research and Development Program of China; 2020YFS0579 Science & Technology Department of Sichuan Province; ZYYC23015 West China Hospital, Sichuan University; 2023TQ0230 China Postdoctoral Science Foundation

0 (locked nucleic acid)
0 (Oligonucleotides)
0 (DNA, Single-Stranded)
9007-49-2 (DNA)

Date Created: 20240313 Date Completed: 20240424 Latest Revision: 20240426

20240426

PMC11040154

10.1093/nar/gkae176

38477342