Inhibition of CRISPR-Cas12a trans-cleavage by lead (II)-induced G-quadruplex and its analytical application

In this work, the inhibition of clustered regularly interspaced short palindromic repeats (CRISPR) - CRISPR associated protein (Cas) trans-cleavage by Pb²⁺-induced G-quadruplex has been firstly explored to detect Pb²⁺ in tea beverage and milk. In absence of Pb²⁺, the Na⁺-induced G-quadruplex can be cleaved by CRISPR-Cas12a. In contrast, Pb²⁺ can competitively combine with G-quadruplex, resulting in its conformational changes and resistance to trans-cleavage of CRISPR-Cas12a. Therefore, the fluorescence resonance energy transfer can happen. Pb²⁺ can be detected in a linear range from 100 nM to 5 µM with a lowest detection limit of 2.6 nM and a relative standard deviation of 4.32%. In summary, this work not only provides a new method for Pb²⁺ detection based on its induced G-quadruplex inhibition on CRISPR-Cas12a trans-cleavage, but also broadens the application of CRISPR-Cas system for heavy metal analysis in the field of food safety.