DNA isolation: Cancer genomic DNA was extracted from formalin-fixed paraffin-embedded tissue using a MagCore® Genomic DNA FFPE One-Step Kit (RBC Bioscience, Taiwan). The quality was quantified using a DeNovix DS-11 Spectrophotometer (DeNovix, Wilmington, DE, USA) and a QuantiFluo® ONE dsDNA System (Promega, Madison, WI, USA).
The assay generates a library of 207 gene-specific amplicons and targets ~2800 clinically relevant mutations.
Sequencing: The products were analyzed via next-generation sequencing (NGS) using an Illumina platform, MiSeq Dx.
Data analysis: the analysis of NGS data was performed using the GALAXY platform (usegal-axy.org, accessed on). Sequencing reads (FASTQ files) were aligned to the human reference genome hg19 using the Bowtie2 tool. Variant calling was performed using the Varscan2 tool. The parameters used for the analysis were minimum allele frequency—0.05, minimum quality—20, and minimum coverage ×80. All variants were annotated with ANNOVAR (https://wannovar.wglab.org , accessed on 2 May 2022). The results were visualized using the R Bioconductor package by Maftools (http://bioconductor.org/ , accessed on 2 May 2022). More details have been described in previous study [18 (link)].
The assay generates a library of 207 gene-specific amplicons and targets ~2800 clinically relevant mutations.
Sequencing: The products were analyzed via next-generation sequencing (NGS) using an Illumina platform, MiSeq Dx.
Data analysis: the analysis of NGS data was performed using the GALAXY platform (usegal-axy.org, accessed on). Sequencing reads (FASTQ files) were aligned to the human reference genome hg19 using the Bowtie2 tool. Variant calling was performed using the Varscan2 tool. The parameters used for the analysis were minimum allele frequency—0.05, minimum quality—20, and minimum coverage ×80. All variants were annotated with ANNOVAR (
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