DRAGEN TSO 500 and TSO 500 ctDNA Analysis Software
  • DRAGEN TSO 500 and TSO 500 ctDNA Analysis Software
  • DRAGEN TSO 500 Guides
    • DRAGEN TSO 500 v2.6
      • Introduction to DRAGEN TSO 500 Analysis Software v2.6.x
      • Getting Started
        • Installation of 2.6.0 on Standalone DRAGEN Server
        • Installation of 2.6.1 on Standalone DRAGEN Server
        • Getting Started on Illumina Connected Analytics
        • Installation of NovaSeq 6000Dx TSO 500 Analysis Application
      • Run Planning
        • Sample Sheet Introduction
        • Sample Sheet Requirements
        • Sample Sheet Creation in BaseSpace Run Planning tool
        • NovaSeq 6000Dx Run Set Up
        • Sample Sheet Templates
      • Launching Analysis
        • Analysis Launch on Standalone DRAGEN Server
          • Command-Line Options
          • Run on Multiple DRAGEN Servers
        • Analysis Launch on ICA
          • Auto-Launch of DRAGEN TSO 500 Analysis on ICA
          • Manual Launch of DRAGEN TSO 500 Analysis on ICA
      • Analysis Output
        • DNA Output
        • RNA Output
        • Combined Variant Output
        • Metrics Output
          • DNA Expanded Metrics
          • RNA Expanded Metrics
          • HRD Metrics Report
        • Coverage Reports
        • Block List
      • Analysis Methods
        • FastQ Generation
        • DNA Analysis Methods
        • RNA Analysis Methods
        • Quality Control
        • Performance Testing
      • Troubleshooting
    • DRAGEN TSO 500 v2.5
      • Introduction to DRAGEN TSO 500 Analysis Software v2.5.x
      • Getting Started
        • Installation of 2.5.3 on Standalone DRAGEN Server
        • Installation of v2.5.4 on Standalone DRAGEN Server
        • Getting Started on Illumina Connected Analytics
      • Run Planning
        • Sample Sheet Introduction
        • Sample Sheet Requirements
        • Sample Sheet Creation in BaseSpace Run Planning tool
        • Sample Sheet Templates
      • Launching Analysis
        • Analysis Launch on Standalone DRAGEN Server
          • Command-Line Options
          • Run on Multiple DRAGEN Servers
        • Analysis Launch on ICA
          • Auto-Launch of DRAGEN TSO 500 Analysis on ICA
          • Manual Launch of DRAGEN TSO 500 Analysis on ICA
      • Analysis Output
        • DNA Output
        • RNA Output
        • Combined Variant Output
        • Metrics Output
          • DNA Expanded Metrics
          • RNA Expanded Metrics
          • HRD Metrics Report
          • Block List
        • Analysis Methods
          • FASTQ Generation
          • DNA Analysis Methods
          • RNA Analysis Methods
          • Quality Control
          • Performance Testing
      • Troubleshooting
  • DRAGEN TSO 500 ctDNA Guides
    • DRAGEN TSO 500 ctDNA v2.6
      • Introduction to DRAGEN TSO 500 ctDNA Analysis Software v2.6
      • Getting Started
        • Installation of TSO 500 ctDNA v2.6.0 on Standalone DRAGEN Server
        • Installation of TSO 500 ctDNA v2.6.2 on Standalone DRAGEN Server
        • Getting Started on Illumina Connected Analytics
        • Installation of NovaSeq 6000Dx TSO 500 ctDNA Analysis Application
      • Run Planning
        • Sample Sheet Introduction
        • Sample Sheet Requirements
        • Sample Sheet Creation in BaseSpace Run Planning tool
        • NovaSeq 6000Dx Run Set Up
        • Sample Sheet Templates
      • Launching Analysis
        • Analysis Launch on Standalone DRAGEN Server
          • Command-Line Options
        • Analysis Launch on ICA
          • Auto-Launch of DRAGEN TSO 500 ctDNA Analysis on ICA
          • Manual Launch of DRAGEN TSO 500 ctDNA Analysis on ICA
      • Analysis Output
        • DNA Output
        • Combined Variant Output
        • Metrics Output
          • DNA Expanded Metrics
        • Coverage Reports
        • Block List
      • Analysis Methods
        • FastQ Generation
        • DNA Analysis Methods
        • Quality Control
      • Troubleshooting
  • Reference
    • Revision History
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  • Run QC
  • NextSeq 500/550 or NextSeq 550Dx (RUO)
  • NovaSeq 6000 or NovaSeq 6000Dx (RUO)
  • NextSeq 1000/2000
  • NovaSeq X
  • DNA Sample QC
  • RNA Sample QC

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  1. DRAGEN TSO 500 Guides
  2. DRAGEN TSO 500 v2.6
  3. Analysis Methods

Quality Control

The software calculates several quality control metrics for runs and samples.

Run QC

The Run Metrics section of the metrics output report provides sequencing run quality metrics along with suggested values to determine if they are within an acceptable range. The overall percentage of reads passing filter is compared to a minimum threshold. For Read 1 and Read 2, the average percentage of bases ≥ Q30, which gives a prediction of the probability of an incorrect base call (Q‑score), are also compared to a minimum threshold. The following tables show run metric and quality threshold information for different systems.

The values in the Run Metrics section are listed as NA in the following situations:

  • If the analysis was started from FASTQ files.

  • If the analysis was started from BCL files and the InterOp files are missing or corrupt.

NextSeq 500/550 or NextSeq 550Dx (RUO)

Metric
Description
Recommended Guideline Quality Threshold
Variant Class

PCT_PF_READS (%)

Total percentage of reads passing filter.

≥80.0

All

PCT_Q30_R1 (%)

Percentage of Read 1 reads with quality score ≥ 30.

≥80.0

All

PCT_Q30_R2 (%)

Percentage of Read 2 reads with quality score ≥ 30.

≥80.0

All

NovaSeq 6000 or NovaSeq 6000Dx (RUO)

Metric
Description
Recommended Guideline Quality Threshold
Variant Class

PCT_PF_READS (%)

Total percentage of reads passing filter.

≥55.0

All

PCT_Q30_R1 (%)

Percentage of Read 1 reads with quality score ≥ 30.

≥80.0

All

PCT_Q30_R2 (%)

Percentage of Read 2 reads with quality score ≥ 30.

≥80.0

All

NextSeq 1000/2000

Metric
Description
Recommended Guideline Quality Threshold
Variant Class

PCT_PF_READS (%)

Total percentage of reads passing filter.

≥85.0

All

PCT_Q30_R1 (%)

Percentage of Read 1 reads with quality score ≥ 30.

≥85.0

All

PCT_Q30_R2 (%)

Percentage of Read 2 reads with quality score ≥ 30.

≥85.0

All

NovaSeq X

Metric
Description
Recommended Guideline Quality Threshold
Variant Class

PCT_Q30_R1 (%)

Percentage of Read 1 reads with quality score ≥ 30.

≥85.0

All

PCT_Q30_R2 (%)

Percentage of Read 2 reads with quality score ≥ 30.

≥85.0

All

DNA Sample QC

DRAGEN TruSight Oncology 500 uses QC metrics to assess the validity of analysis for DNA libraries that pass contamination quality control. If the library fails one or more quality metrics, then the corresponding variant type or biomarker is not reported, and the associated QC category in the report header displays FAIL. Additionally, a companion diagnostic result may not be available if it relies on QC passing for one or more of the following QC categories.

DNA library QC results are available in the MetricsOutput.tsv file. Refer to Metrics Output for details.

Metric
Description
Recommended Guideline Quality Threshold
Variant Class

CONTAMINATION_SCORE

The contamination score is based on VAF distribution of SNPs.

≤ 1457

All

MEDIAN_EXON_COVERAGE

Median exon fragment coverage across all exon bases.

≥ 150

Small variant TMB

PCT_EXON_50X

Percent exon bases with 50x fragment coverage.

≥ 90.0

Small variant TMB

MEDIAN_INSERT_SIZE

The median fragment length in the sample.

≥ 70

Small variant TMB

USABLE_MSI_SITES

The number of MSI sites usable for MSI calling.

≥ 40

MSI

MEDIAN_BIN_COUNT_CNV_TARGET

The median raw bin count per CNV target.

≥ 1.0

CNV

RNA Sample QC

The input for RNA Library QC is RNA alignment. Metrics and guideline thresholds can be found in the MetricsOutput.tsv file. Refer to Metrics Output for details.

Metric
Description
Recommended Guideline Quality Threshold
Variant Class

MEDIAN_CV_GENE_500X

The median CV for all genes with median coverage > 500x. Genes with median coverage > 500x are likely to be highly expressed. Higher CV median > 500x indicates an issue with library preparation (poor sample input and/or probes pulldown issue).

Fusion Splice

MEDIAN_INSERT_SIZE

The median fragment length in the sample.

≥ 80

Fusion Splice

TOTAL_ON_TARGET_READS

The total number of reads that map to the target regions.

≥ 9000000

Fusion Splice

GENE_MEDIAN_COVERAGE

The median deduped coverage across all genes in the RNA panel (55 genes).

N/A*

Fusion Splice

  • To avoid failing RNA samples unnecessarily, Illumina does not recommend a universal threshold to determine RNA sample quality. RNA expression varies significantly across tissue types and a small panel size (55 genes), which makes normalization challenging. Tissue-specific thresholds could be considered for normalization.

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Last updated 2 months ago

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