A DNA-Cu nanocluster and exonuclease I integrated label-free reporting system for CRISPR/Cas12a-based SARS-CoV-2 detection with minimized background signals

J Mater Chem B. 2022 Aug 17;10(32):6107-6117. doi: 10.1039/d2tb00857b.

Abstract

CRISPR-driven biosensing is developing rapidly, but current studies mostly adopt dye-labeled ssDNA as the signal reporter, which is costly and unstable. Herein, we developed a label-free and low-background reporter for CRISPR/Cas12a signaling by integrating DNA-templated copper nanoclusters (DNA-CuNCs) and exonuclease I (EXO I). The template of the DNA-CuNCs was rationally designed as a ds-/ss-DNA hybrid, ensuring that after a quick and nonpersistent cut of Cas12a, a majority of the template can be digested by EXO I. Based on this novel reporter, a biosensor termed CRISPR-CNS (cost-effective, nimble, and sensitive copper nanocluster sensor integrating CRISPR) was developed. Due to the high signal-to-background ratio of our proposed reporter, CRISPR-CNS shows excellent performances for nucleic acid detection, yielding a detection limit of 20 copies for SARS-CoV-2 RNA. Considering its facile synthesis, robust fluorescence, effective cost, and good sensitivity, this combination shall serve as a highly potential output for CRISPR-based point-of-care testing.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • COVID-19* / diagnosis
  • CRISPR-Cas Systems / genetics
  • Copper
  • DNA / genetics
  • Exodeoxyribonucleases
  • Humans
  • RNA, Viral
  • SARS-CoV-2* / genetics

Substances

  • RNA, Viral
  • Copper
  • DNA
  • Exodeoxyribonucleases
  • exodeoxyribonuclease I