A CRISPR-Cas12a integrated SERS nanoplatform with chimeric DNA/RNA hairpin guide for ultrasensitive nucleic acid detection

Bohan Yin, Qin Zhang, Xinyue Xia, Chuanqi Li, Willis Kwun Hei Ho, Jiaxiang Yan, Yingying Huang, Honglian Wu, Pui Wang, Changqing Yi, Jianhua Hao, Jianfang Wang, Honglin Chen, Siu Hong Dexter Wong, Mo Yang

Research output: Journal article publicationJournal articleAcademic researchpeer-review

30 Citations (Scopus)

Abstract

Background: CRISPR-Cas12a has been integrated with nanomaterial-based optical techniques, such as surface-enhanced Raman scattering (SERS), to formulate a powerful amplification-free nucleic acid detection system. However, nanomaterials impose steric hindrance to limit the accessibility of CRISPR-Cas12a to the narrow gaps (SERS hot spots) among nanoparticles (NPs) for producing a significant change in signals after nucleic acid detection. Methods: To overcome this restriction, we specifically design chimeric DNA/RNA hairpins (displacers) that can be destabilized by activated CRISPR-Cas12a in the presence of target DNA, liberating excessive RNA that can disintegrate a core–satellite nanocluster via toehold-mediated strand displacement for orchestrating a promising “on-off” nucleic acid biosensor. The core–satellite nanocluster comprises a large gold nanoparticle (AuNP) core surrounded by small AuNPs with Raman tags via DNA hybridization as an ultrabright Raman reporter, and its disassembly leads to a drastic decrease of SERS intensity as signal readouts. We further introduce a magnetic core to the large AuNPs that can facilitate their separation from the disassembled nanostructures to suppress the background for improving detection sensitivity. Results: As a proof-of-concept study, our findings showed that the application of displacers was more effective in decreasing the SERS intensity of the system and attained a better limit of detection (LOD, 10 aM) than that by directly using activated CRISPR-Cas12a, with high selectivity and stability for nucleic acid detection. Introducing magnetic-responsive functionality to our system further improves the LOD to 1 aM. Conclusion: Our work not only offers a platform to sensitively and selectively probe nucleic acids without pre-amplification but also provides new insights into the design of the CRISPR-Cas12a/SERS integrated system to resolve the steric hindrance of nanomaterials for constructing biosensors.

Original languageEnglish
Pages (from-to)5914-5930
Number of pages17
JournalTheranostics
Volume12
Issue number13
DOIs
Publication statusPublished - 8 Aug 2022

Keywords

  • CRISPR-Cas12a
  • gold nanoparticles
  • magnetic manipulation
  • nucleic acid detection
  • surface-enhanced Raman spectroscopy

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Pharmacology, Toxicology and Pharmaceutics (miscellaneous)

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