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Torrent suite

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The Torrent Suite is a software package designed for analyzing and managing data generated by Thermo Fisher Scientific's Torrent sequencing instruments. It provides tools for primary analysis, reporting, and data management.

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Lab products found in correlation

Ion reporter, by Thermo Fisher Scientific (7 mentions) Ion chef system, by Thermo Fisher Scientific (6 mentions) Ion ampliseq custom panel, by Thermo Fisher Scientific (4 mentions) Ampliseqrna analysis plugin, by Thermo Fisher Scientific (4 mentions) Recoverall total nucleic acid isolation kit, by Thermo Fisher Scientific (4 mentions) Ion ampliseq designer, by Thermo Fisher Scientific (3 mentions) Ion torrent proton platform, by Thermo Fisher Scientific (3 mentions) Qubit rna high sensitivity assay, by Thermo Fisher Scientific (3 mentions) Qubit fluorometer, by Thermo Fisher Scientific (3 mentions) Ion s5 system, by Thermo Fisher Scientific (3 mentions) Ion ampliseq exome rdy panel, by Thermo Fisher Scientific (3 mentions) 540 chip chef kit, by Thermo Fisher Scientific (3 mentions) Torrent suite software, by Thermo Fisher Scientific (2 mentions) Ion sphere particle, by Thermo Fisher Scientific (2 mentions) Ampure xp system, by Thermo Fisher Scientific (2 mentions) Qubit dsdna hs assay kit reagent, by Thermo Fisher Scientific (2 mentions) Ion chef, by Thermo Fisher Scientific (2 mentions) Qubit 3.0 fluorometer, by Thermo Fisher Scientific (2 mentions) Ion proton, by Thermo Fisher Scientific (2 mentions) Allprep dna rna mini kit, by Qiagen (2 mentions)

Spelling variants of this product

Ion Torrent Suite Software (22 citations) Torrent Suite v4.0 (11 citations) Torrent suite software v3.6.2 (7 citations) Torrent suite v. 5.10.1 (6 citations) Torrent Suite software version 5.10.1 (4 citations) Torrent Suite™ Server (3 citations) Torrent Suite v (2 citations) Torrent Suite, version 4.2 (2 citations) Torrent Suite Software 4.0 (2 citations) Torrent Suite 3.4 (2 citations)

78 protocols using torrent suite

1

Variant Analysis of SARS-CoV-2 Genomes

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Sequenced reads were aligned with the Wuhan-Hu-1 Reference Genome (NC_045512.2) on the Torrent Suite v. 5.12.2 (Life Technologies, Carlsbad, California, USA). Plugins were used as follows by order coverage analysis (v5.12.0.0) and Variant Caller (v.5.12.0.4), both with default parameters and COVID19AnnotateSnpEff (v1.3.0.2; Life Technologies, Carlsbad, California, USA). To predict the effect of a base substitution, a plugin specifically developed for SARS-CoV-2. VCF files generated by Torrent Suite (Life Technologies, Carlsbad, California, USA). Variant caller was filtered to remove variants with read depths less than 1000 and ion torrent quality scores less than 400 to keep reliable variants only (S1 Table). The filtered variants were used for sample clustering with Maximum Likelihood Tree in Molecular Evolutionary Genetics Analysis (MEGA, https://www.megasoftware.net) software. The consensus for each SARS-CoV-2 genome sequence were then submitted to the GISAID [9 ] under the accession numbers of EPI_ISL_1402423 to EPI_ISL_1477049 (available for registered users) and NCBI under the accession numbers GI:2021275696 to GI:2021275839 (or MW853559.1 to MW853569.1) [10 ] databases (Table 1 and S1 File).
Ayubov M.S., Buriev Z.T., Mirzakhmedov M.K., Yusupov A.N., Usmanov D.E., Shermatov S.E., Ubaydullaeva K.A, & Abdurakhmonov I.Y. (2022). Profiling of the most reliable mutations from sequenced SARS-CoV-2 genomes scattered in Uzbekistan. PLoS ONE, 17(3), e0266417.
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2

Mitochondrial DNA Sequence Analysis

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DNA sequences were reconstructed by pooling all barcoded libraries to respective samples and trimming adapter sequences 20 bases from the 3' and 5' end, using Torrent Suite™ software (Applied Biosystems, CA, USA). Sequences were formatted to the human mitochondrial genome by alignment to the revised Cambridge Reference Sequence (rCRS) [22, 23] . Sequence variants, SNPs, insertions and deletions (INDELs) were reported using the Ion PGM™ System: Torrent Variant Caller (Applied Biosystems, CA, USA) plugin as variant caller files. Binary alignment map files of aligned sequences and variants were inspected using Integrative Genomics Viewer (Broad Institute, MA, USA) [24, 25] . Variants were imported into MitoTool, a third party online software designed with PhyloTree Build 17, and used for assignment of mitochondrial haplogroups [26, 27] . HaploGrep 2 (v2.1.0) was used as a secondary confirmation of haplogroups [28] . A minimum arbitrary threshold of 10X coverage reads was used to call mitochondrial variants and a threshold of 0.05 was set for point heteroplasmy detection. In line with forensic convention, length heteroplasmy was reported to the most dominant allele of all detected sequences [29] .
Wai K.T., Barash M, & Gunn P. (2018). Performance of the Early Access AmpliSeq™ Mitochondrial Panel with degraded DNA samples using the Ion Torrent™ platform. Electrophoresis, 39(21).
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3

Deep Sequencing of HCV Variants

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The nucleotide sequences obtained from each sample were aligned with reference sequences (H77: NC004102 and Con1: AJ238800) obtained from the Los Alamos HCV sequence database using Torrent Suite™ software (Applied Biosystems). For analysis of sequence variants and coverage, we used the plugins Variant Caller and Coverage Analyzer, respectively, both available in the Torrent Suite software. The quality of called bases of Ion Torrent PGM was based on an algorithmic phred scale (Q20) with 99% of accuracy. The frequency of indels through alignment to the reference demonstrated that Ion Torrent PGM reads had 1.5 indels per 100 bases (1.72 indels per read) [21] . So, to identify variants, we used low stringency parameters, with a threshold of 2%, minimum quality of 100, minimum coverage of 2, minimum coverage on either strand of 1, maximum strand bias of 0.95 and minimum relative read quality of 6.5.
For the NS3-4A region, we analysed resistanceassociated mutations at amino acid positions 36 The average of the coverages obtained for the analysis of variants of the NS3-4A regions, NS5A and NS5B were 387.2x, 516.7x and 165.7x, respectively.
Gaspareto K.V., Ribeiro R.M., de Mello Malta F., Gomes-Gouvêa M.S., Muto N.H., Romano C.M., Mendes-Correa M.C., Carrilho F.J., Sabino E.C, & Rebello Pinho J.R. (2016). Resistance-associated variants in HCV subtypes 1a and 1b detected by Ion Torrent sequencing platform. Antiviral therapy, 21(8).
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4

Mapping Protein-Mitochondrial RNA Interactions

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To identify physiological interactions between proteins and mitochondrial RNAs cross linking immunoprecipitation (CLIP) was carried out essentially as previously described (Dennerlein et al, 2010 (link)). To generate a more comprehensive data set, the final PCR products were prepared for Ion Torrent sequencing, following manufacturer's instructions, rather than subcloning into pCR4-TOPO. Sequence data for 100 000–190 000 reads was collected, aligned to mtDNA as a reference sequence using the Torrent Suite software on the Ion Torrent server. The alignments were then viewed using IGV (integrative genomics viewer) and presented against a circular depiction of human mtDNA, using a log scale to indicate the number of reads per RNA site.
Hornig-Do H.T., Montanari A., Rozanska A., Tuppen H.A., Almalki A.A., Abg-Kamaludin D.P., Frontali L., Francisci S., Lightowlers R.N, & Chrzanowska-Lightowlers Z.M. (2014). Human mitochondrial leucyl tRNA synthetase can suppress non cognate pathogenic mt-tRNA mutations. EMBO Molecular Medicine, 6(2), 183-193.
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5

Optimized Variant Calling in Ion Torrent Sequencing

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Data runs were initially processed using the Ion Torrent platform-specific pipeline software, Torrent Suite, to generate sequence reads, trim adapter sequences, filter, and remove poor signal-profile reads. Initial variant calling from the Ion AmpliSeq sequencing data was generated using Ion Torrent cloud-based pipeline Ion Reporter v5.0. We used cut-off values greater than 10% of the variant frequency and more than ×200 coverage to detect true variants in accordance with previous reports and our own experience. To eliminate erroneous base calling, the second filter was employed by visually examining the mutations using CLC Genomics Workbench version 9.5.1 (Qiagen, Hilden, Germany), as well as by filtering out possible strand-specific errors.
Fujita S., Masago K, & Yatabe Y. (2019). Biopsy of palliative lesions following radiotherapy. BJR Open, 1(1), 20180025.
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6

Ion Torrent Sequencing of Fast-TT-COLD-PCR Amplicons

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Following fast-TT-COLD-PCR, mutation-enriched amplicons from each temperature window were processed using the standard barcoding and library preparation protocol for Ion Torrent sequencing. Libraries with ligated Ion Torrent adapters were assessed for DNA quality and quantity on Agilent bio-analyzer, and then pooled together into a single tube prior to Ion Torrent sequencing. An emulsion PCR containing Ion Sphere Particles (ISPs) and libraries with ligated Ion Torrent adapters were performed to optimally amplify amplicons bound to ISPss using Ion Torrent’s One Touch2™ instrument and Ion PGM Template OT2 200 Kit.
Following enrichment of template-positive Ion Sphere Particles, templates were sequenced with the Ion Torrent Machine (PGM using the 318 chip) per manufacturer’s protocol. Data analysis including base calling, read filtering, and demultiplexing were performed using the Torrent Suite software (version 3.4.2 or higher) from Ion Torrent. Variants were detected using the variant Caller plugin from Torrent Suite. Reads for each non-reference bases at the nucletotide of interest as well as the adjacent nucleotides were recorded and plotted in a noise plot to distinguish the true mutation versus background noise. Unfiltered NGS data (BAM files) were also loaded into Integrative Genome Viewer 2.3 (IGV, Broad Institute) (33 (link)) using human genome hg19 as reference.
Castellanos-Rizaldos E., Richardson K., Lin R., Wu G, & Makrigiorgos G.M. (2014). SINGLE-TUBE, HIGHLY PARALLEL MUTATION ENRICHMENT IN CANCER GENE PANELS USING TEMPERATURE-TOLERANT-COLD-PCR. Clinical chemistry, 61(1), 267-277.
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7

Genome-wide Copy Number Analysis

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Sequencing alignment and coverage analyses were performed using Torrent Suite version 5.0.2 (Ion Torrent, Carlsbad, CA). Genome-wide copy number alterations were first called from aligned, non-PCR-duplicate reads using the QDNASeq R package (version 1.6.1) [69 (link)]. Segmented copy-number events were identified using bin-level corrected, median- and control-normalized read counts using the circular binary segmentation algorithm implemented by the DNACopy (1.44.0) R package, and final segment- and bin-level copy-number values were used for subsequent analyses as described (see Supplementary Methods). Focal CNAs were defined as CNAs 1.5–20 Mb long with a log2(CopyNumberRatio) ≥ 0.2.
Hovelson D.H., Liu C.J., Wang Y., Kang Q., Henderson J., Gursky A., Brockman S., Ramnath N., Krauss J.C., Talpaz M., Kandarpa M., Chugh R., Tuck M., Herman K., Grasso C.S., Quist M.J., Feng F.Y., Haakenson C., Langmore J., Kamberov E., Tesmer T., Husain H., Lonigro R.J., Robinson D., Smith D.C., Alva A.S., Hussain M.H., Chinnaiyan A.M., Tewari M., Mills R.E., Morgan T.M, & Tomlins S.A. (2017). Rapid, ultra low coverage copy number profiling of cell-free DNA as a precision oncology screening strategy. Oncotarget, 8(52), 89848-89866.
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8

Iron-Associated Gene Panel Sequencing

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Sequencing of genomic DNA using an iron-associated gene panel, covering 39 genes and 11 promoters associated with iron overload or anemia, was performed as previously described [15 ]. Samples were sequenced to an average of 400-fold coverage using an Ion Torrent PGM, with post run QC/QA filtering performed using Torrent Suite (v3.6; Life Technologies). Sequences were mapped to the Human Genome (HG19) and variants identified using the Torrent Variant Caller (version 3.6.59049 with Germ Line - Low Stringency configuration). Identified variants were annotated using the wANNOVAR tool [16 (link)], and candidate mutations confirmed by Sanger sequencing.
McDonald C.J., Rishi G., Secondes E.S., Ostini L., Wallace D.F., Crawford D.H., Sia H., Clark P, & Subramaniam V.N. (2018). Evaluation of a bone morphogenetic protein 6 variant as a cause of iron loading. Human Genomics, 12, 23.
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9

Comprehensive Genetic Mutation Profiling

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Sequencing data were analyzed using Torrent Suite (Version 3.2.0) (Life Technologies). Frequency (%) of all of the common KRAS (codons 12 and 13), BRAF (V600E) and EGFR (T790M and L858R) gene point mutations and those at five randomly picked base positions with nucleotide G or C in amplified regions of chromosomes 7 and 12 were calculated. The C:G >T:A mutations include KRAS G12D (GGT>GAT), G12S (GGT>AGT), G13D (GGC>GAC), G13S (GGC>AGC), and EGFR T790M (ACG>ATG). The non-C:G >T:A mutations include KRAS G12A (GGT>GCT), G12C (GGT>TGT), G12R (GGT>CGT), G13A (GGC>GCC), G13C (GGC>TGC), G13R (GGC>CGC), and BRAF V600E (GTG>GAG).
Chen G., Mosier S., Gocke C.D., Lin M.T, & Eshleman J.R. (2014). Cytosine Deamination is a Major Cause of Baseline Noise in Next Generation Sequencing. Molecular diagnosis & therapy, 18(5), 587-593.
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10

Comprehensive Mutation Analysis Pipeline

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Sequencing data of the three targeted genes (KRAS, BRAF, and EGFR) were analyzed using Torrent Suite (version 3.2.0; Life Technologies). All other genes except KRAS, BRAF, and EGFR were masked for this analysis in Torrent Variant Caller using specific browser extensible data files and in Ion Reporter using filters. Mutations were identified and annotated through Torrent Variant Caller (version 3.2.45211) and Ion Reporter (version 1.2) Figure 1. Additional analysis using visual inspection of the binary sequence alignment/map file on the Broad Institute's (Boston, MA) Integrative Genomics Viewer (IGV) was implemented (early in the validation process) after we found that Torrent Variant Caller and Ion Reporter missed a case with the most common EGFR point mutation in a lung cancer (L858R). IGV was used to determine the coverage of each specific exon and to display the number of reads of the variants. The Torrent Variant Caller plugin called the nucleotide variants along with their Hg19 genome position. Ion Reporter identified nucleotide variants with integrated annotations and provided a COSMIC ID if available. Annotation was confirmed by direct inspection of the nucleotide sequences and amino acid sequences. The annotations from these three pathways were compared.
Lin M.T., Mosier S.L., Thiess M., Beierl K.F., Debeljak M., Tseng L.H., Chen G., Yegnasubramanian S., Ho H., Cope L., Wheelan S.J., Gocke C.D, & Eshleman J.R. (2014). Clinical Validation of KRAS, BRAF, and EGFR Mutation Detection Using Next-Generation Sequencing. American journal of clinical pathology, 141(6), 856-866.
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