Amplification-free RNA detection with CRISPR–Cas13
Hajime Shinoda, Yuya Taguchi, Ryoya Nakagawa, Asami Makino, Sae Okazaki, Masahiro Nakano, Yukiko Muramoto, Chiharu Takahashi, Ikuko Takahashi, Jun Ando, Takeshi Noda, Osamu Nureki, Hiroshi Nishimasu, Rikiya Watanabe
Abstract
CRISPR-based nucleic-acid detection is an emerging technology for molecular diagnostics. However, these methods generally require several hours and could cause amplification errors, due to the pre-amplification of target nucleic acids to enhance the detection sensitivity. Here, we developed a platform that allows "CRISPR-based amplification-free digital RNA detection (SATORI)", by combining CRISPR-Cas13-based RNA detection and microchamber-array technologies. SATORI detected single-stranded RNA targets with maximal sensitivity of ~10 fM in <5 min, with high specificity. Furthermore, the simultaneous use of multiple different guide RNAs enhanced the sensitivity, thereby enabling the detection of the SARS-CoV-2 N-gene RNA at ~5 fM levels. Therefore, we hope SATORI will serve as a powerful class of accurate and rapid diagnostics.