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. 2023 Aug 6;12(8):1017.
doi: 10.3390/pathogens12081017.

An ARMS-Multiplex PCR Targeting SARS-CoV-2 Omicron Sub-Variants

Petros Bozidis  1 , Eleni Petridi  1 , Konstantina Gartzonika  1
Affiliations

An ARMS-Multiplex PCR Targeting SARS-CoV-2 Omicron Sub-Variants

Petros Bozidis et al. Pathogens. .

Abstract

As of November 2021, the SARS-CoV-2 Omicron variant had made its appearance, gradually replacing the predominant Delta variant. Since its emergence, the Omicron variant has been continuously evolving through more than 500 strains, most of which belong to five sub-variants known as BA.1, BA.2, BA.3, BA.4, and BA.5. The aim of this study was to develop a multiplex polymerase chain reaction (PCR) that will be able to distinguish the basic sub-variants of Omicron in a rapid and specific way. Full genome sequences of Omicron strains with high frequency and wide geographical distribution were retrieved by the NCBI Virus and ENA databases. These sequences were compared to each other in order to locate single nucleotide polymorphisms common to all strains of the same sub-variant. These polymorphisms should also be capable of distinguishing Omicron sub-variants not only from each other but from previously circulating variants of SARS-CoV-2 as well. Thus, specific primers targeting characteristic polymorphisms of the four Omicron main branches BA.1, BA.2, BA.4, and BA.5 were designed according to the principles of the amplification refractory mutation system (ARMS) and with the ability to react under multiplex PCR conditions. According to our results, the ARMS-multiplex PCR could successfully distinguish all Omicron sub-variants that carry the corresponding mutations.

Keywords: ARMS; BA.1; BA.2; BA.4; BA.5; Multiplex PCR; Omicron; SARS-CoV-2; variants.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Sequences were retrieved from the NCBI Virus and ENA databases. Selected sequences were confirmed by the Pangolin COVID-19 Lineage assigner and BLAST tools. Only sequences that were verified by 100% and 99%, respectively, were selected for nucleotide comparison within the same sub-variant group.
Figure 2
Figure 2
Agarose gel electrophoresis (1.5% w/v) of ARMS–multiplex PCR products generated by the amplification of four cDNAs corresponding to the four Omicron (BA.1, BA.2, BA.4, and BA.5) sub-variants. A 100 bp DNA ladder was used to confirm the correct length of the anticipated amplicons. The size of each amplicon is indicated on the left. NC, negative control (without DNA).
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