Topological barrier to Cas12a activation by circular DNA nanostructures facilitates autocatalysis and transforms DNA/RNA sensing

Abstract Control of CRISPR/Cas12a trans -cleavage is crucial for biosensor development. Here, we show that small circular DNA nanostructures which partially match guide RNA sequences only minimally activate Cas12a ribonucleoproteins. However, linearizing these structures restores activation. Building on this finding, an Autocatalytic Cas12a Circular DNA Amplification Reaction (AutoCAR) system is established which allows a single nucleic acid target to activate multiple ribonucleoproteins, and greatly increases the achievable reporter cleavage rates per target. A rate-equation-based model explains the observed near-exponential rate trends. Autocatalysis is also sustained with DNA nanostructures modified with…

• NF
  National Foundation for Medical Research and Innovation
• AC
  ARC Centre for Nanoscale BioPhotonics

  Award: CE14010003
• CI
  Cancer Institute NSW

  Award: 2019/CDF1013
• UO
  University of New South Wales
• UO
  University of Technology Sydney
• FP
  Fondation pour la Recherche Médicale
• MR
  Medical Research Council
• NH
  National Health and Medical Research Council

  Awards: GNT1195742, 2019/CDF1013, GNT1181889