Multiplex PCR panel for SARS-CoV-2 amplicon sequencing
The ARTIC panel consists of two primer pools that amplify SARS-CoV-2 viral genomes from complex samples for research. The sequencing data can elucidate the presence of mutations consistent with known variants or identify new variants in research studies.
xGen NGS—made for COVID-19 research.
Using next generation sequencing (NGS) to analyze SARS-CoV-2 can help researchers understand COVID-19. IDT, in partnership with the ARTIC Network, is offering their latest design of the ARTIC SARS-Co-V-2 Amplicon Panel to support global availability of this important research tool.
The ARTIC network—a group of researchers who are developing a system for processing different virus samples to generate public health information—has developed primer sequences, experimental recommendations, and bioinformatic resources to facilitate NGS analysis of the SARS-CoV-2 genome. Their primer design pipeline (PrimalSeq), previously deployed for other infectious disease sequencing, was especially useful early in the SARS-COV2 pandemic .
ARTIC provides a detailed experimental protocol featuring a set of primers for amplicon sequencing to quickly sequence SARS-CoV-2 . They also maintain a resource that alerts researchers to updates in the design . An adaptative protocol that allows for sequencing on Illumina sequencers have been developed by others .
The sequence of the SARS-CoV-2 genome can enable research in many areas, including:
The ARTIC SARS-CoV-2 Amplicon Panel workflow includes cDNA synthesis, amplicon generation, and cleanup. The final amplicons average 400 bp require 2 x 250 bp read lengths on either the NovaSeq™ or MiSeq™ sequencers (Illumina).
Table 1. Specifications for the ARTIC SARS-CoV-2 Amplicon Panel
|Recommended Ct value||Coverage||Contiguous amplicons||Coordinates covered|
Figure 1. The ARTIC SARS-CoV-2 Amplicon Panel’s genome coverage for Omicron variants. Two nasopharyngeal swab samples (Cts 28.4 and 13.6) were subjected to the protocol using the ARTIC v4.1 primer pool to create amplicons, then the xGen DNA Library Prep MC Library Prep Kit to adapt the amplicons to create an NGS library. Libraries were sequenced on a MiSeq™ (Illumina) with 2 x 250 bp PE reads. Data was down sampled to 100,000 reads. Alignment was then performed using the Burrows-Wheeler Alignment (BWA) Tool  and a consensus file uploaded to Pangolin for lineage calling. High genomic coverage for the BA.1 lineage was observed (IGV screenshot above) indicating the compatibility of the panel with Omicron variants.
Table 2. Sequencing metrics for ARTIC v4.1 libraries created from nasopharyngeal swabs.
|Sample||Ct||Reads aligned||On-target||Coverage uniformity||% Genome >10X||Lineage|
Sequencing metrics for NGS libraries originating from nasopharyngeal swabs. Libraries were created with the ARTIC v4.1 primer pool and the xGen DNA Library Prep MC Library Prep Kit. Libraries were sequenced on a MiSeq™ (Illumina) with 2 x 250 bp PE reads. Data was down sampled to 100,000 reads. Lineages were called with Pangolin.
*RUO—For research use only. Not for use in diagnostic procedures. Unless otherwise agreed to in writing, IDT does not intend for these products to be used in clinical applications and does not warrant their fitness or suitability for any clinical diagnostic use. Purchaser is solely responsible for all decisions regarding the use of these products and any associated regulatory or legal obligations.