iScience
Volume 24, Issue 11, 19 November 2021, 103329
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Article
Endonuclease-based genotyping of the RBM as a method to track the emergence or evolution of SARS-CoV-2 variants

https://doi.org/10.1016/j.isci.2021.103329Get rights and content
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Highlights

  • Mutations in SARS-CoV-2 RBM are important for virus adaptation

  • We developed a genotyping assay on the RBM coding sequence

  • The genotyping is based on the use of a mismatch-specific endonuclease

  • The test can help monitor the emergence and circulation of VOCs

Summary

Since the beginning of the COVID-19 pandemics, variants have emerged. Some of them display increased transmissibility and/or resistance to immune response. Most of the mutations involved in the functional adaptation are found in the receptor-binding motif (RBM), close to the interface with the receptor ACE2. We thus developed a fast molecular assay to detect mutations in the RBM coding sequence. After amplification, the amplicon is heat-denatured and hybridized with an amplicon of reference. The presence of a mutation can be detected using a mismatch-specific endonuclease and the cleavage pattern is analyzed by capillary electrophoresis. The method was validated on RNA of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants produced in vitro before being implemented for clinical samples. The assay showed 97.8% sensitivity and 97.8% specificity. The procedure can be set up for high-throughput identification of the presence of mutations and serve as a first-line screening to select the samples for full genome sequencing.

Subject areas

Virology
Methodology in biological sciences

Data and code availability

  • All data reported in this paper will be shared by the lead contact upon request.

  • This paper does not report original code.

  • Any additional information required to reanalyze the data reported in this paper is available from the lead contact upon request.

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Lead contact