A CRISPR-Cas12a-based specific enhancer for more sensitive detection of SARS-CoV-2 infection

Huang, W.; Yu, L.; Wen, D.; Wei, D.; Sun, Y.; Zhao, H.; Ye, Y.; Chen, W.; Zhu, Y.; Wang, L.; Wu, W.; Zhao, Q.; Xu, Y.; Gu, D.; Nie, G.; Zhu, D.; Guo, Z.; Ma, X.; Niu, L.; Huang, Y.; Liu, Y.; Peng, B.; Zhang, R.; Zhang, X.; Li, D.; Yang, G.; Liu, L.; Zhou, Y.; Wang, Y.; Hou, T.; Gao, Q.; Li, W.; Chen, S.; Hu, X.; Han, M.; Zheng, H.; Wen, J.; Cai, Z.; Song, F.; Zhao, G.; Wang, J.

High Ct-values falling in the grey zone are frequently encountered in SARS-CoV-2 detection by real-time reverse transcription PCR (rRT-PCR) and have brought urgent challenges in diagnosis of samples with low viral load. Based on the single-stranded DNA reporter trans-cleavage activity by Cas12a upon target DNA recognition, we create a Specific Enhancer for detection of PCR-amplified Nucleic Acids (SENA) to confirm SARS-CoV-2 detection through specifically targeting its rRT-PCR amplicons. SENA is highly sensitive, with its limit of detection being at least 2 copies/reaction lower than that of the corresponding rRT-PCR, and highly specific, which identifies both false-negative and false-positive cases in clinic applications. SENA provides effective confirmation for nucleic acid amplification-based molecular diagnosis, and may immediately eliminate the uncertainty problems of rRT-PCR in SARS-CoV-2 clinic detection.