Targeted genome sequencing (TS) is an effective NGS method that focuses on the interesting regions of the genome and enables the rapid detection of genetic variations implicated in diseases. Through selective enrichment of the targeted genomic regions, TS can profile a panel of genes or biomarkers with known clinical relevance to discover powerful mutations, rather than screening the whole genome via large-scale DNA sequencing (whole genome sequencing, WGS). TS shows better accuracy and sensitivity in identifying targeted variations owing to greater sequencing depth at the same sequencing cost and data burden when compared with WGS or whole exome-sequencing (WES)1. Thus, it allows high confidence to identify mutations with low variant allele frequencies (VAFs), especially for low-quality and fragmented clinical DNA samples. Clinical targeted sequencing has revolutionized tumor surveillance and diagnosis and facilitated the development of precision oncology2,3. TruSight™ Oncology 500 (TSO500, Illumina) is a comprehensive target-sequencing panel that covers more than eight cancer types, 523 cancer-related genes (1.94 Mb) to identify relevant genomic variants and signatures in a single assay (https://www.illumina.com/products/by-brand/trusight-oncology-500.html). We can obtain multiple variant information from formalin-fixed paraffin-embedded (FFPE) tissue specimens through the TSO500 assay4. These mutations include small nucleotide variants (SNVs), InDels, copy number variations (CNVs), splice variants and gene fusions. In addition, it accurately measures key current immuno-oncology biomarkers: microsatellite instability (MSI) and tumor mutational burden (TMB). TSO500 workflow allows variants detection with high specificity and sensitivity in FFPE samples, especially for mutants with low VAFs via tactfully suppressing technical noise and excluding germline variants. TSO500 assay has displayed exemplary performance in profiling TMB, MSI, and various gene variants in different tumor types5–7.
Many high-throughput sequencers are compatible with targeted genome sequencing, and have been widely applied in scientific studies and clinical diagnosis. NextSeq550 system (Illumina), a flexible sequencing platform with multiple read length and output configurations, has been commonly used in WES, transcriptome, as well as targeted sequencing8,9. It was reported that the TSO500 panel performed well for variant screening of myeloid neoplasm coupled with the NextSeq550 platform10. The high sensitivity, specificity, precision, and accuracy demonstrated the clinical applicability of this assay for SNVs and InDels/duplications detection. Similarly, on the NovaSeq 6000 platform (Illumina), the TSO500 panel also showed excellent performances and good reproducibility in identifying multiple DNA/RNA variant types and molecular signatures from FFPE tissue specimens11–13. GenoLab M (GeneMind LTD., China), a novel high-throughput sequencing platform, utilizes the well-recognized sequencing-by-synthesis (SBS) techniques and reversible termination approaches to deliver confident base-calling accuracy and high yield of error-free reads. Moreover, GenoLab M is highly compatible with Illumina Library Preparation Kits, and 2X150 bp paired-end sequencing mode is available. Previously, the performance metric of GenoLab M on transcriptome and LncRNA sequencing for three model species (mouse, bean, and human) has been compared with NovaSeq 6000 platform. This budding platform shows comparable data quality in detecting gene expression, alternatively spliced (AS) events, single nucleotide polymorphism (SNP), and InDel14.
The study aimed to evaluate whether the GenoLab M platform can accurately detect mutations in tumor standard sample. The OncoSpan FFPE (HD832) sample was captured by the TSO500 panel through hybrid-capture-based target enrichment, and sequenced by NovaSeq 6000, NextSeq 550, and GenoLab M platforms. We compared the performances of three sequencing platforms.