Issue 6, 2021

Amplified detection of nucleic acids and proteins using an isothermal proximity CRISPR Cas12a assay

Abstract

Herein, we describe an isothermal proximity CRISPR Cas12a assay that harnesses the target-induced indiscrimitive single-stranded DNase activity of Cas12a for the quantitative profiling of gene expression at the mRNA level and detection of proteins with high sensitivity and specificity. The target recognition is achieved through proximity binding rather than recognition by CRISPR RNA (crRNA), which allows for flexible assay design. A binding-induced primer extension reaction is used to generate a predesigned CRISPR-targetable sequence as a barcode for further signal amplification. Through this dual amplification protocol, we were able to detect as low as 1 fM target nucleic acid and 100 fM target protein isothermally. The practical applicability of this assay was successfully demonstrated for the temporal profiling of interleukin-6 gene expression during allergen-mediated mast cell activation.

Graphical abstract: Amplified detection of nucleic acids and proteins using an isothermal proximity CRISPR Cas12a assay

Supplementary files

Article information

Article type
Edge Article
Submitted
06 nov 2020
Accepted
16 dec 2020
First published
04 jan 2021
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY-NC license

Chem. Sci., 2021,12, 2133-2137

Amplified detection of nucleic acids and proteins using an isothermal proximity CRISPR Cas12a assay

Y. Li, H. Mansour, C. J. F. Watson, Y. Tang, A. J. MacNeil and F. Li, Chem. Sci., 2021, 12, 2133 DOI: 10.1039/D0SC06113A

This article is licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported Licence. You can use material from this article in other publications, without requesting further permission from the RSC, provided that the correct acknowledgement is given and it is not used for commercial purposes.

To request permission to reproduce material from this article in a commercial publication, please go to the Copyright Clearance Center request page.

If you are an author contributing to an RSC publication, you do not need to request permission provided correct acknowledgement is given.

If you are the author of this article, you do not need to request permission to reproduce figures and diagrams provided correct acknowledgement is given. If you want to reproduce the whole article in a third-party commercial publication (excluding your thesis/dissertation for which permission is not required) please go to the Copyright Clearance Center request page.

Read more about how to correctly acknowledge RSC content.

Social activity

Spotlight

Advertisements