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Issue 14, 2020
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Direct observation of the formation of a CRISPR–Cas12a R-loop complex at the single-molecule level

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Abstract

Here, we develop an optical tweezers-based single-molecule manipulation assay to detect the formation of an R-loop complex in the Cas12a system and characterize its thermodynamic stability. We found that the formation of the R-loop complex induces a two-step unfolding of a DNA hairpin containing the target sequence, the non-target sequence binds loosely to Cas12a and can be easily released from the complex, and the Nuc domain of Cas12a plays key roles in target binding and R-loop formation.

Graphical abstract: Direct observation of the formation of a CRISPR–Cas12a R-loop complex at the single-molecule level

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Article information


Submitted
24 Oct 2019
Accepted
10 Jan 2020
First published
14 Jan 2020

Chem. Commun., 2020,56, 2123-2126
Article type
Communication

Direct observation of the formation of a CRISPR–Cas12a R-loop complex at the single-molecule level

Y. Cui, Y. Tang, M. Liang, Q. Ji, Y. Zeng, H. Chen, J. Lan, P. Jin, L. Wang, G. Song and J. Lou, Chem. Commun., 2020, 56, 2123
DOI: 10.1039/C9CC08325A

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