RNA was purified using a mirVana miRNA isolation kit (Thermo Fisher Scientific). RNA libraries were generated using an Ion Total RNA-Seq kit v2 (Life Technologies) according to the manufacturer’s instructions. The RNA libraries were then processed for emulsion PCR using an Ion OneTouchTM system and an Ion OneTouch 200 Template kit v3 (Life Technologies). Template-positive Ion SphereTM particles were enriched and purified for the sequencing reaction with an Ion OneTouchTM ES system (Life Technologies). The template-positive Ion SphereTM particles were then applied on Ion PI™ chips (Life Technologies) and a high-throughput sequencing reaction was carried out using an Ion Proton™ semiconductor sequencer (Life Technologies). All of the sequencing data were mapped on a human reference genome sequence (GRCh37/hg19) using the Torrent Suite software program (Life Technologies). The expression analysis for each sample was imported into the CLC Genomics Workbench software program (CLC Bio, Aarhus, Denmark) and the significance of the differences among the samples was determined by an unpaired t-test. The gene ontology (GO) analysis was performed using the MetaCore software.
Torrent suite software program
Manufactured by Thermo Fisher Scientific
Sourced in United States
Torrent Suite software program is a comprehensive bioinformatics software suite developed by Thermo Fisher Scientific. It provides a platform for the analysis and management of next-generation sequencing data. The software's core function is to facilitate the processing, alignment, and interpretation of sequencing data generated by Thermo Fisher's Ion Torrent sequencing instruments.
Automatically generated - may contain errors
Lab products found in correlation
Ion onetouch es system, by Thermo Fisher Scientific (2 mentions)
Ion total rna seq kit v2, by Thermo Fisher Scientific (2 mentions)
Ion pi chip, by Thermo Fisher Scientific (2 mentions)
Ion onetouch system, by Thermo Fisher Scientific (2 mentions)
Clc genomics workbench software program, by Qiagen (2 mentions)
Ion proton semiconductor sequencer, by Thermo Fisher Scientific (2 mentions)
Ion onetouch 200 template kit v3, by Thermo Fisher Scientific (2 mentions)
Mirvana mirna isolation kit, by Thermo Fisher Scientific (1 mentions)
3 protocols using torrent suite software program
1
RNA Sequencing for Transcriptome Analysis
2
RNA-Seq Protocol for Differential Gene Expression
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Basically, the analysis was performed using methods previously reported by our group23 (link). RNA libraries were generated using an Ion Total RNA-Seq Kit v2 (Thermo Fisher Scientific) according to the manufacturer’s instructions. Emulsion PCR was carried out with an Ion OneTouchTM system and an Ion OneTouch 200 Template Kit v3 (Thermo Fisher Scientific). Template-positive Ion Sphere™ particles were enriched and purified for the sequencing reaction with an Ion OneTouch™ ES system (Thermo Fisher Scientific). The template-positive Ion Sphere™ Particles were loaded onto Ion PI™ Chips (Thermo Fisher Scientific) and for high throughput sequencing with an Ion Proton™ Semiconductor sequencer (Thermo Fisher Scientific). Sequencing data were mapped on a human reference genome sequence (GRCh38/hg38) using the Torrent Suite software program (Life Technologies). The expression analysis was performed in the CLC Genomics Workbench software program (CLC bio, Aarhus, Denmark), and differences among the samples were determined using an unpaired Student’s t-test. The gene list describing fold change and p-value was uploaded to the MetaCore software (Clarivate Analytics, PA, USA, URL; https://portal.genego.com/ , version 6.33.69110.), and then pathway analysis was performed.
3
Genetic Variant Analysis Pipeline
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All of the sequencing data were mapped on a human reference genome sequence (GRCh37/hg19) using the Torrent Suite Software program (Life technologies, Carlsbad, CA, USA). The genetic variants were then detected by a Torrent Variant Caller plug-in for the software program (Life technologies, Carlsbad, CA, USA). In this program, alleles with frequencies of (the percentage of reads which possessed a variant) >10%, with a coverage (the number of reads which had a variant) of >5 and a quality score of >15 were regarded as significant variants. The variant information for each sample was imported into the CLC Genomics Workbench software system (CLC bio, Aarhus, Denmark), and Fisher's exact test was performed to determine the significance of the differences among the samples. Strand bias was defined according to the following numerical formula: strand bias=max (VpCm, VmCp)/VpCm+VmCp (Cp, the number of reads from the plus direction in the known sequence; Cm, the number of reads from the minus direction in the known sequence; Vp, the number of reads from the plus direction in the variant sequence; Vm, the number of reads from the minus direction in the variant sequence).
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