Harnessing noncanonical trans-cleavage characteristics of Cas12 and Cas13a to enhance CRISPR-based diagnostics
Year: 2024
Type: article
Abstract: Cas12 and Cas13 are extensively utilized in molecular diagnostics for their trans-cleavage activities, yet their activation characteristics remain partially understood. Here, we conduct an in-depth investigation of Cas12a, Cas12f1, and Cas13a, uncovering the characteristics of their trans-DNase and trans-RNase activities with noncanonical activators. Our findings reveal that DNA can serve as a direct target for CRISPR-Cas13a, markedly increasing the detection sensitivity for single-base mismatches. Moreover, the trans-cleavage activities of Cas12a and Cas13a can be activated by diverse RNA:DNA and RNA:RNA duplexes, respectively, indicating that the presence of stem-loop structures in crRNAs is not essential for their activation. Notably, Cas12f1, unlike Cas12a, exhibits intrinsic RNase activity independently of activation. Leveraging these insights, we have improved the accuracy of a dual-gene target detection approach that employs the CRISPR-Cas12f1 and Cas13a systems. Our research advances the understanding of the noncanonical activation characteristics of Cas12 and Cas13a, contributing to the field of CRISPR-based diagnostics.
Source: Communications Biology
Authors: Qing Wu, Zhengfei Yi, Haoran Li, Guoxin Han, Jianyong Du, Jingwei Xiong, Keping Hu, Hai Gao
Institutions: Chinese Academy of Medical Sciences & Peking Union Medical College, Institute of Medicinal Plant Development, Peking University, State Key Laboratory of Natural and Biomimetic Drugs, PLA 306 Hospital, Chinese People's Liberation Army, University of Health and Rehabilitation Sciences, Shanghai Medical College of Fudan University, Fudan University
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,890
Awards: G1927020422, G852789696
OpenAlex ID: W4403347796