Genestudio s5 sequencer

Libraries were prepared with the Ion AmpliSeq Library Kit Plus according to the manufacturer’s instruction using Ion AmpliSeq SARS-CoV-2 RNA custom primers panel (ID: 05280253, ThermoFisher Scientific). The RNA library preparation included reverse transcription using SuperScript VILO cDNA Synthesis Kit (ThermoFisher Scientific) with subsequently 16–21 cycles of PCR amplification on Ion Chef. Next-generation sequencing (NGS) reactions were run on Ion Torrent GeneStudio S5 sequencer.
Sequence alignments to the SARS-CoV-2 isolate Wuhan-Hu-1, complete genome (NCBI nucleotide collection, accession number: NC_045512) (Wu et al., 2020 (link)), was performed within the Torrent Server of Ion Torrent S5 sequencer using default settings. The aligned reads were utilized for both reference-guided assemblies. Assembly was performed using the Iterative Refinement Meta-Assembler (IRMA) v.0.6.1 (Shepard et al., 2016 (link)) that produced a consensus sequence for each sample using a >50% cutoff for calling single-nucleotide polymorphisms. IRMA utilizes multiple steps of alignment, variant calling, and consensus building by capitalizing on multiple allele frequency confidence intervals and read depth. Aligned reads were validated through the Integrative Genomics Viewer (IGV) v.2.5.3 (Thorvaldsdóttir et al., 2013 (link)).

Libraries were prepared with the Ion AmpliSeq Library Kit Plus according to the manufacturer’s instruction using Ion AmpliSeq SARS-CoV-2 RNA custom primers panel (ID: 05280253, Thermo Fisher Scientific). The RNA library preparation included reverse transcription using SuperScript VILO cDNA Synthesis Kit (Thermo Fisher Scientific) with subsequently 16–21 cycles of PCR amplification on the Ion Chef platform. Next-generation sequencing (NGS) reactions were run on Ion Torrent GeneStudio S5 sequencer.
Sequence alignments to the SARS-CoV-2 isolate Wuhan-Hu-1, complete genome (NCBI nucleotide collection, accession number: NC_045512.2) (Wu et al., 2020 (link)), was performed within the Torrent Server of Ion Torrent S5 sequencer using default settings. The aligned reads were utilized for both reference-guided assemblies. Assembly was performed using the Iterative Refinement Meta-Assembler (IRMA) v.0.6.1 (Shepard et al., 2016 (link)) that produced a consensus sequence for each sample using a >50% cutoff for calling single-nucleotide polymorphisms. IRMA utilizes multiple steps of alignment, variant calling, and consensus building by capitalizing on multiple allele frequency confidence intervals and read depth. Aligned reads were validated through the Integrative Genomics Viewer (IGV) v.2.9.4 (Thorvaldsdóttir et al., 2013 (link)).