A qualitative RT-PCR assay for the specific identification of the SARS-CoV-2 B.1.1.529 (Omicron) Variant of Concern

J Clin Virol. 2022 Jul:152:105191. doi: 10.1016/j.jcv.2022.105191. Epub 2022 May 24.


Objectives: The aim of this study was to develop a RT-PCR assay for the specific detection of the SARS-CoV-2 Omicron Variant of Concern (VOC) as a rapid alternative to sequencing.

Methods: A RT-PCR was designed in silico and then validated using characterised clinical samples containing Omicron (both BA.1 and BA.2 lineages) and the Omicron synthetic RNA genome. As negative controls, SARS-CoV-2 positive clinical samples collected in May 2020, and synthetic RNA genomes of the isolate Wuhan Hu-1 and of the Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Kappa (B.1.617.1), Iota (B.1.526), Epsilon (B.1.429) and Delta (B.1.617.2) SARS-CoV-2 VOC were used.

Results: Experiments performed using as templates the synthetic RNA genomes demonstrate the high specificity of the PCR-method for the SARS-CoV-2 Omicron. Despite the synthetic RNAs were used at high copy numbers, specific signal was mainly detected with the Omicron synthetic genome. Only a non-specific late signal was detected using the Alpha variant genome, but these results were considered negligible as Alpha VOC has been replaced by the Delta and it is not circulating anymore in the world. Using our method, we confirmed the presence of Omicron on clinical samples containing this variant but not of other SARS-CoV-2 lineages. The method is highly sensitive and can detect up to 1 cp of the Omicron virus per µl.

Conclusions: The method presented here, in combination with other methods in use for detection of SARS-CoV-2, can be used for an early identification of Omicron.

Keywords: B.1.1.529; Omicron; RT-PCR; SARS-CoV-2; Specific identification.

MeSH terms

  • COVID-19* / diagnosis
  • Humans
  • RNA, Viral / analysis
  • Reverse Transcriptase Polymerase Chain Reaction
  • SARS-CoV-2* / genetics
  • Sensitivity and Specificity


  • RNA, Viral

Supplementary concepts

  • SARS-CoV-2 variants