Vigilant: An Engineered VirD2-Cas9 Complex for Lateral Flow Assay-Based Detection of SARS-CoV2

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Article
Authors
Marsic, Tin
Ali, Zahir cc
Tehseen, Muhammad cc
Mahas, Ahmed cc
Hamdan, Samir cc
Mahfouz, Magdy M. cc
KAUST Department
Bioengineering
Biological and Environmental Science and Engineering (BESE) Division
Bioscience Program
Center for Desert Agriculture
Laboratory for Genome Engineering
Laboratory for Genome Engineering and Synthetic Biology, Division of Biological Sciences, King Abdullah University of Science and Technology (KAUST), Thuwal 23955-6900, Saudi Arabia
Laboratory of DNA Replication and Recombination
Plant Science
Date
2021-04-12
Online Publication Date
2021-04-12
Print Publication Date
2021-04-28
Embargo End Date
2022-04-12
Submitted Date
2021-02-12
Permanent link to this record
http://hdl.handle.net/10754/668815

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Abstract
Rapid, sensitive, and specific point-of-care testing for pathogens is crucial for disease control. Lateral flow assays (LFAs) have been employed for nucleic acid detection, but they have limited sensitivity and specificity. Here, we used a fusion of catalytically inactive SpCas9 endonuclease and VirD2 relaxase for sensitive, specific nucleic acid detection by LFA. In this assay, the target nucleic acid is amplified with biotinylated oligos. VirD2-dCas9 specifically binds the target sequence via dCas9 and covalently binds to a FAM-tagged oligonucleotide via VirD2. The biotin label and FAM tag are detected by a commercially available LFA. We coupled this system, named Vigilant (<u>Vi</u>rD2-dCas9 <u>g</u>u<u>i</u>ded and <u>L</u>FA-coupled <u>n</u>ucleic acid <u>t</u>est), to reverse transcription-recombinase polymerase amplification to detect SARS-CoV2 in clinical samples. Vigilant exhibited a limit of detection of 2.5 copies/μL, comparable to CRISPR-based systems, and showed no cross-reactivity with SARS-CoV1 or MERS. Vigilant offers an easy-to-use, rapid, cost-effective, and robust detection platform for SARS-CoV2.
Citation
Marsic, T., Ali, Z., Tehseen, M., Mahas, A., Hamdan, S., & Mahfouz, M. (2021). Vigilant: An Engineered VirD2-Cas9 Complex for Lateral Flow Assay-Based Detection of SARS-CoV2. Nano Letters. doi:10.1021/acs.nanolett.1c00612
Sponsors
We would like to thank Mohammad Alarawi for providing the RNA of SARS-CoV-2 clinical samples. We also thank members of the genome engineering and synthetic biology laboratory for insightful discussions and technical support. This work was supported, in part, by the Smart Health Initiative at KAUST and the IAF grant from the KAUST IED to M.M.
Publisher
American Chemical Society (ACS)
Journal
Nano Letters
DOI
10.1021/acs.nanolett.1c00612
PubMed ID
33844549
Additional Links
https://pubs.acs.org/doi/10.1021/acs.nanolett.1c00612
Collections
Articles; Biological and Environmental Science and Engineering (BESE) Division; Bioscience Program; Center for Desert Agriculture
This document is the Accepted Manuscript version of a Published Work that appeared in final form in Nano Letters, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://pubs.acs.org/doi/10.1021/acs.nanolett.1c00612.
Except where otherwise noted, this item's license is described as This document is the Accepted Manuscript version of a Published Work that appeared in final form in Nano Letters, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see https://pubs.acs.org/doi/10.1021/acs.nanolett.1c00612.