Systematic Noise Filtering
The DRAGEN systematic noise filter is available in somatic mode, and can be used to reduce false positive calls by accounting for site-specific noise. The filter can be used in T/N mode, but is especially useful for T/O runs where a matched normal is not available.
To use the DRAGEN systematic noise filter you need to:
| 1. | Generate Systematic Noise BED Files. or Use Prebuilt Systematic Noise BED Files |
| 2. | Run DRAGEN Somatic Variant Caller with a Systematic Noise Filter |
Generate Systematic Noise BED Files
The systematic noise BED should be generated from normal samples. It is recommended to build systematic noise files that are library prep, sequencing system, and panel specific. During step 2 the most appropriate noise file can be used to remove noise when analyzing somatic samples. For noise generation it is recommended to use approximately 50 normal samples.
| 1. | To generate VCF output per normal sample, run DRAGEN in somatic tumor-only on approximately 50 normal samples at a time with --vc-detect-systematic-noise set to true. The purpose of this step is to detect noise present in normal samples. This step will ignore any filters, so there is no need to specify filters during this step. |
Only use --vc-detect-systematic-noise to generate noise files. The option is not intended for analyzing tumor samples.
| 2. | Run DRAGEN in somatic mode by specifying inputs with --tumor-fastq1, --tumor-fastq2, or --tumor-bam-input. |
| 3. | Build the BED file using the VCFs and the following options. |
You can also build systematic noise BED files in the cloud using the BaseSpace Sequence Hub DRAGEN CNV Baseline Builder App.
|
Option |
Description |
|---|---|
|
--vc-systematic-noise-raw-input-list |
List of input VCFs. Enter one VCF per line. |
|
--vc-systematic-noise-germline-vaf-threshold |
Minimum VAF to remove potential germlines from systematic noise file building. Variants with a VAF greater than the threshold are not considered systematic noise. The default is none. The default is unspecified, which indicates that all variants are used. If using small panels, the recommended threshold is 0.3. |
|
--vc-systematic-noise-use-germline-tag |
Use DRAGEN internal germline tagging to remove potential germlines. Mutually exclusive with --vc-systematic-noise-germline-vaf-threshold. The default is false. If using WGS or WES, the recommended setting is true. |
|
--vc-systematic-noise-method |
Method to calculate the systematic noise level (noise allele frequency) across samples. Enter mean to calculate the average noise allele frequency, max to calculate the maximum, or aggregate to calculate total alleles / total depth per locus across samples. The default is mean. If using WGS the recommended setting is max. If using WES, the recommended setting is aggregate. If using small panels for higher sensitivity, the recommended setting is mean or aggregate. |
Use Prebuilt Systematic Noise BED Files
The following prebuilt systematic noise files for WGS and WES are available for download on the Illumina DRAGEN Bio-IT Platform support site page.
|
Prebuilt Systematic Noise File |
Comment |
Number of Normal Samples |
|---|---|---|
|
WGS_hg38_v1.0_systematic_noise.bed.gz |
WGS hg38 |
28 Samples. The samples are a mixture of the Illumina DNA PCR-free kit sequenced on the NovaSeq 6000, Illumina DNA PCR-free kit sequenced on the PCR-free HiSeq X, and the TruSeq DNA Nano kit HiSeq X. |
|
WGS_hs37d5_v1.0_systematic_noise.bed.gz |
WGS hs37d5 |
31 Samples. The samples are a mixture of the Illumina DNA PCR-free kit sequenced on the NovaSeq 6000, Illumina DNA PCR-free kit sequenced on theHiSeq X, and the TruSeq DNA Nano sequenced on the HiSeq X. |
|
WGS_hg19_v1.0_systematic_noise.bed.gz |
WGS hg19 |
31 Samples. The samples are a mixture of the Illumina DNA PCR-free kit sequenced on the NovaSeq 6000, Illumina DNA PCR-free kit sequenced on theHiSeq X, and the TruSeq DNA Nano sequenced on the HiSeq X. |
|
WES_Nextera_IDT_hg38_v1.0_systematic_noise.bed.gz |
Nextera library prep; IDT exome; hg38 |
47 Samples |
|
WES_Nextera_IDT_hs37d5_v1.0_systematic_noise.bed.gz |
Nextera library prep; IDT exome; hs37d5 |
47 Samples |
|
WES_Nextera_IDT_hg19_v1.0_systematic_noise.bed.gz |
Nextera library prep; IDT exome; hg19 |
47 Samples |
|
WES_TruSeq_IDT_hg38_v1.0_systematic_noise.bed.gz |
TruSeq library prep; IDT exome; hg38 |
53 Samples |
|
WES_TruSeq_IDT_hs37d5_v1.0_systematic_noise.bed.gz |
TruSeq library prep; IDT exome; hs37d5 |
53 Samples |
|
WES_TruSeq_IDT_hg19_v1.0_systematic_noise.bed.gz |
TruSeq library prep; IDT exome; hg19 |
53 Samples |
Run DRAGEN Somatic Variant Caller with a Systematic Noise Filter
When DRAGEN is used in somatic mode, you can specify a BED file with site-specific noise level to filter out sequencing / systematic noise. The site-specific noise level is used to calculate a Phred-scaled AQ score. If the AQ score is smaller than the defined threshold, the variant is filtered as systematic noise.
The following systematic noise command line options are available:
|
Option |
Description |
|---|---|
|
--vc-systematic-noise— |
Specifies a systematic noise BED file. If a somatic variant does not pass the AQ threshold, the variant is marked as systematic_noise in the FILTER column of the output VCF. |
|
--vc-systematic-noise-filter-threshold |
Set the AQ threshold. By default the threshold value for tumor-normal is 10 and 60 for tumor-only. |
