Definition of CRISPR Cas12a Trans-Cleavage Units to Facilitate CRISPR Diagnostics
Hailong Lv, Jian Wang, Jian Wang, Jian Zhang, Yijian Chen, Lei Yin, Dian Jin, Dayong Gu, Huailong Zhao, Yong Xu, Jin Wang, Jin Wang
Abstract
The CRISPR diagnostic (CRISPR-Dx) technology that employs the trans -cleavage activities has shown great potential in diagnostic sensitivity, specificity, convenience, and portability, and has been recognized as the next-generation diagnostic methods. However, due to the lack of standardized definition of Cas trans -cleavage enzymatic units, it is difficult to standardize the present CRISPR-Dx systems, which have undoubtedly impeded the development of the CRISPR-Dx industry. To solve the problem, we here first systematically optimized the reaction systems for Cas12a, and then defined its trans -cleavage units ( trans U), which we believe will be of great importance and interest to researchers in both molecular diagnostic industry and basic research. Moreover, a simple protocol was provided to facilitate a step-by-step measurement of the Cas12a trans U, which can also act as a reference for the definition of the trans U for other Cas proteins.